> Chemicals & Drugs > Amino Acid > Cyclic Peptides

Cyclic Peptides

Q: What are the unique properties of cyclic peptides?

Cyclic peptides have a circular structure formed by a covalent bond between the amino and carboxyl termini. This confers several advantageous properties, including increased stability, resistance to proteolytic degradation, and improved cell permeability compared to linear peptides. The cyclic structure also gives cyclic peptides unique physicochemical and pharmacological activities, making them valuable for various therapeutic applications.

Q: How are cyclic peptides found and produced?

Cyclic peptides can be found naturally in microorganisms, plants, and animals. They can also be synthetically produced through chemical synthesis or recombinant methods. The ability to engineer cyclic peptides has expanded their potential applications in areas like drug development, diagnostics, and biotechnology.

Q: What are some common applications of cyclic peptides?

Cyclic peptides have a wide range of applications due to their unique properties. They are being researched and utilized in the development of novel drugs, particularly for oncology, antimicrobial, and anti-inflammatory indications. Cyclic peptides are also used in the design of diagnostice tools and as reagents in biotechnological processes.

Q: How can PubCompare.ai assist in cyclic peptide research?

PubCompare.ai's AI-driven platform can accelerate cyclic peptide research in several ways. First, it allows researchers to screen protocol literature more efficiently, helping them quickly locate relevant protocols from published literature, pre-prints, and patents. Second, the platform's AI-powered analysis can pinpointe critical insights, enabling researchers to identify the most effective protocols related to cyclic peptides for their specific research goals. This helps ensure reproducibility and accuracy by highlighting key differences in protocol effectiveness, allowing researchers to choose the best option.

Q: What are some common challenges in working with cyclic peptides?

One of the main challenges in working with cyclic peptides is their relatively poor solubility and cell permeability compared to linear peptides. This can make it difficult to formulate and deliver cyclic peptides effectively. Additionally, the synthesis of cyclic peptides can be more complex and time-consuming than linear peptides. Researchers may also face challenges in consistently producing cyclic peptides with the desired purity and tytpical properties.

Cyclic Peptides are a class of peptides with a circular structure formed by a covalent bond between the amino and carboxyl termini.
These molecules exhibit unique physicochemical properties and pharmacological activities, making them valuable in various therapeutic applications.
Cyclic Peptides are found naturally in microorganisms, plants, and animals, and can also be synthetically produced.
Their cyclic structure confers increased stability, resistance to proteolytic degradation, and improved cell permeability compared to linear peptides.
Researching Cyclic Peptides can lead to the development of novel drugs, diagnostics, and biotechnological tools.
PubCompare.ai's AI-driven platform can accelerate Cyclic Peptide research by helping scientists easily locate relevant protocols from literature, pre-prints, and patents, and leverage AI-powered comparisons to identify the best protocols and products.

Most cited protocols related to «Cyclic Peptides»

Extending mMass for Cyclic Peptides

Cited 28 times

On a search for a software tool satisfying as many as possible of the criteria discussed above, one of the authors (TN) identified the software mMass to be the most suitable basis for further development. mMass is a portable and cross-platform open-source software for mass spectra processing, written mainly in Python [37] (link), which already offered a GUI, spectrum manipulation capabilities, the possibility to enter and save peptide sequences and do in-silico fragmentation and spectrum annotation. However, entering and fragmentation of peptides was limited to linear sequences and proteinogenic amino acids, and a user-editable monomer database as well as several fragmentation options important for cyclic peptides were missing. Since mMass is open-source, the software could be modified to meet all of the criteria described above.
First of all, a monomer library editor has been designed, giving the user the possibility to conveniently enter, edit and save monomers needed for the composition of peptide sequences. The library has been filled with all monomers compiled from the NORINE database [16] , facilitating its use for new users and allowing for easy addition of own variants. Furthermore, the monomer editor allows for the definition of possible neutral losses from individual monomers.
Secondly, the sequence organization and handling has been extended to enable the composition of peptides containing non-proteinogenic amino acids. For this purpose, an additional sequence editor has been designed that allows the convenient composition of peptides by typing or dragging and dropping monomers from the monomer library right into the editor. In addition, a peptide can be set as being linear or cyclic.
Finally, the fragmentation module has been improved to handle all possible known fragmentation pathways of cyclic peptides, to allow for the loss of custom neutrals, and to allow for multiple neutral losses from one fragment.
To assess the capabilities of the resulting software package, several cyanobacterial natural product MS² spectra have been annotated, and the annotations have been compared to those made by NRP-Annotation.

Niedermeyer T.H, & Strohalm M. (2012). mMass as a Software Tool for the Annotation of Cyclic Peptide Tandem Mass Spectra. PLoS ONE, 7(9), e44913.

Full text: Click here

Publication 2012

Amino Acids Cyanobacteria Cyclic Peptides DNA Library Mass Spectrometry Natural Products Peptides Python

Structural Refinement of Cyclic Peptides Bound to Mouse Pgp

Cited 17 times

To achieve improved structures of cyclic peptides bound to mouse Pgp, the final drug-free mouse Pgp structure was first refined into the 4.4 Å QZ59-RRR and 4.35 Å QZ59-SSS datasets without drug (rigid body, TLS, NCS, group-, and individual- B-factors). Anomalous difference Fourier maps were then calculated (CNSv1.3) from model phases of the refined structures to pinpoint the location of the selenium atoms in the datasets. Regularized molecules of the RRR- and SSS-cyclic peptides were then manually docked into the structures (in both possible orientations), placing the selenium atoms in anomalous difference Fourier density. The docked structures were then subjected to another round of refinement using cyclic peptide parameter files with the exception that rigid body refinement was omitted. To examine the orientation of the drugs, refined structures were used to calculate Fo-Fc difference maps, and the vicinity of the drug was carefully inspected. As was originally discovered, the “upper” QZ59-SSS molecule was significantly disordered since Fo-Fc density was absent for two vertices of the triangular drug molecule. These atoms were subsequently removed from the final version of the structure.

Li J., Jaimes K.F, & Aller S.G. (2013). Refined structures of mouse P-glycoprotein. Protein Science : A Publication of the Protein Society, 23(1), 34-46.

Publication 2013

3'-(1-butylphosphoryl)adenosine cyclic-tris-(R)-valineselenazole cyclic-tris-(S)-valineselenazole Cyclic Peptides Human Body Mice, House Microtubule-Associated Proteins Muscle Rigidity Pharmaceutical Preparations Selenium

Spectral Networks for Peptide Identification

Cited 15 times

The concept of spectral networks^{37 (link)} (also known as molecular networks^{44 (link)} in the field of natural products) was introduced to reveal spectra of related peptides within a proteomic dataset without knowing what these peptides are. While these networks were first introduced for linear peptides, they were later generalized to cyclic peptides and metabolites^{44 (link),61 (link)}. Nodes in a molecular network correspond to spectra, while edges connect spectra that are generated from related metabolites (e.g., metabolites differing by a single variation). The variations that are captured by molecular networks help to infer mutations, modifications (such as oxidation and acetylation), or adducts (such as sodium and potassium).
DEREPLICATOR+ constructs the molecular network of all spectra and selects connected components with at least one identified spectrum. Using the MSM corresponding to this spectrum, it annotates variants of identified metabolites. Supplementary Figure 10 shows the molecular network of the chalcolmycin family identified by DEREPLICATOR+.

Mohimani H., Gurevich A., Shlemov A., Mikheenko A., Korobeynikov A., Cao L., Shcherbin E., Nothias L.F., Dorrestein P.C, & Pevzner P.A. (2018). Dereplication of microbial metabolites through database search of mass spectra. Nature Communications, 9, 4035.

Full text: Click here

Publication 2018

Acetylation Cyclic Peptides Mutation Natural Products Peptides Potassium Sodium

Computational Pruning of Protein Designs

Cited 14 times

Typically, the number of designs that can be created in silico exceeds the number that can be produced and examined experimentally. We therefore used Rosetta to prune the list of designs, by one of two methods. For design consisting of canonical amino acids, Rosetta’s fragment-based ab initio algorithm^{32 (link)} was utilized to predict a design’s structure given its amino acid sequence, and to determine whether the target structure was a unique minimum in the conformational energy landscape. Disulfide bonds were not allowed to form during these simulations; the designed disulfide bonds are intended to stabilize the folded conformation rather than direct protein folding. Designs which incorporate short stretches of D-amino acids were also validated using Rosetta’s fragment-based ab initio algorithm; the amino acid sequences of designs, with all D-amino acids mutated to glycine, were provided as input, and we allowed Rosetta to generate on the order of 30,000 predicted structures as output. Unlike the standard ab initio protocol, we did not use secondary structure predictions in fragment picking. Additionally, the length of small and large fragments was set to 4 and 6 amino acid residues, instead of the default 3 and 9; we found that this produced better sampling for peptides. After conformational sampling, the D-amino acid positions were changed to their original identities, and rescored. A small modification to the ab initio algorithm permitted it to build a terminal peptide bond for the N-C cyclic designs during the full-atom refinement stages of the structure prediction. Those designs that showed no sampling near the design conformation, or for which the design conformation was not the unique, lowest-energy conformation, were discarded.
Since fragment-based methods are poorly suited to the prediction of structures with large amounts of D-amino acid content, such as NC_cH_LH_R_D1, we developed a new, fragment-free algorithm for validation of these topologies. This algorithm, which we call “simple_cycpep_predict”, uses the same GenKIC-based sampling approach used to build backbones for design, with additional steps of filtering solutions based on disulfide geometry, optimizing sidechain rotamers, and gradient-descent energy minimization. Because the search space is vast, even with the constraints imposed by the N-C cyclic geometry and the disulfide bond(s), we further biased the search by setting mainchain torsion values for residues in the middle of the helices to helical values (a Gaussian distribution centred on phi=−61°, psi=−41° for the α_R helix and on phi=+61°, psi=+41° for the α_L helix); this is analogous to the biased sampling obtained by fragment-based methods, in which sequences with high helix propensity are sampled primarily with helical fragments. As with ab initio validation, designs showing poor sampling near the design conformation or poor energy landscapes were discarded.

Bhardwaj G., Mulligan V.K., Bahl C.D., Gilmore J.M., Harvey P.J., Cheneval O., Buchko G.W., Pulavarti S.V., Kaas Q., Eletsky A., Huang P.S., Johnsen W.A., Greisen P., Rocklin G.J., Song Y., Linsky T.W., Watkins A., Rettie S.A., Xu X., Carter L.P., Bonneau R., Olson J.M., Coutsias E., Correnti C.E., Szyperski T., Craik D.J, & Baker D. (2016). Accurate de novo design of hyperstable constrained peptides. Nature, 538(7625), 329-335.

Publication 2016

Amino Acids Amino Acid Sequence Cyclic Peptides Disulfides Glycine Helix (Snails) Peptides Strains Vertebral Column

High-Resolution Structure of P-glycoprotein

Cited 13 times

X-ray diffraction data were collected at 100 K at either the Stanford Synchrotron Radiation Laboratory (SSRL; BL11-1) or the Canadian Light Source (CLS; 08ID-1). Fluorescence scans were taken on P-gp–cyclopeptide co-crystals to maximize the anomalous signal contribution from the incorporated selenium (Table 1 ▶). All diffraction data were processed with MOSFLM (Battye et al., 2011 ▶ ) and reduced with SCALA (Evans, 2006 ▶ ) within the CCP4 suite of programs (Winn et al., 2011 ▶ ). In the case of QZ-Ala, the data from three isomorphous crystals were scaled together to maximize the completeness (Table 1 ▶). The 3.4 Å resolution structure of P-gp was initially solved by molecular replacement (MR) with Phaser (McCoy et al., 2007 ▶ ) using a previously determined P-gp structure (PDB entry 4ksc; Ward et al., 2013 ▶ ) as a search model with no modifications. Commensurate with the improved resolution, the new electron-density features guided adjustments of our model when compared with the same more ‘open’ crystal form that we reported in 2013 (Ward et al., 2013 ▶ ) and are summarized in Supplementary Fig. S2. Residues 30–32 were located in the electron density, and resulted in a subsequent shift in the registration of residues in the first helix (residues 30–43) preceding TM1. Amendments were made to the topology of intracellular helix 1 (IH1; residues 154–168), extracellular loop 3 (ECL3; residues 318–338) and a portion of TM6 leading into the first NBD (residues 358–387). Within NBD1, residues 398–404, 424–427, 520–526 and 597–602 were rebuilt. Elbow helix 2 (EH2) was rebuilt from residues 689 to 708. A registry issue was amended from ECL4 (residue 738) to TM8 (residue 760) and another that constitutes segments of TM9, ECL5 and the beginning of TM10 (residues 826–855). The topology of IH3 was adjusted (residues 795–806), as was ECL6 (residues 961–967) and a portion of TM12 (residues 972–984). Further modifications were made in the region leading into and contributing to NBD2 (residues 1010–1028, 1042–1047, 1129–1137 and 1165–1172). Residues 1272 and 1273 were also located in the electron-density maps at the C-terminus. As for all structures of P-gp determined to date, the ‘linker’ region (residues 627–688) was not located in the electron density. Many of the structural adjustments are in general agreement with the recent corrections (Li et al., 2014 ▶ ) made to the model of the more ‘closed’ conformation of P-gp first reported in 2009 (Aller et al., 2009 ▶ ). During the refinement process, the model underwent rigid-body and restrained positional refinement, with H atoms applied in their riding positions, using phenix.refine (Afonine et al., 2012 ▶ ) against a maximum-likelihood target function with grouped B factors, secondary-structure restraints, reference-model restraints and TLS. Rounds of refinement were interspersed with manual inspection and correction against σ_A-weighted electron-density maps in Coot (Emsley et al., 2010 ▶ ) and improvements to model geometry and stereochemistry were monitored using MolProbity (Chen et al., 2010 ▶ ). Subsequent cyclopeptide co-crystal structures were solved by either MR or rigid-body refinement using the refined 3.4 Å resolution model with residues from TM4 (218–243) and EH2 (689–694) removed to avoid biasing their placement within the electron-density maps. These structures were then refined in a similar fashion to the 3.4 Å resolution structure with an additional round of positional refinement with ligand B factors set to the Wilson B value. Ligand description dictionaries were determined using phenix.elbow (Adams et al., 2010 ▶ ) and the crystallographic positions of the incorporated seleniums were validated using anomalous scattering methods. The refined structures were judged to have excellent geometry as determined by MolProbity (Chen et al., 2010 ▶ ). The resulting refinement statistics are listed in Table 1 ▶.

Szewczyk P., Tao H., McGrath A.P., Villaluz M., Rees S.D., Lee S.C., Doshi R., Urbatsch I.L., Zhang Q, & Chang G. (2015). Snapshots of ligand entry, malleable binding and induced helical movement in P-glycoprotein. Acta Crystallographica Section D: Biological Crystallography, 71(Pt 3), 732-741.

Full text: Click here

Publication 2015

Complement Factor B Crystallography Cyclic Peptides Elbow Electrons Fluorescence Helix (Snails) Human Body Ligands Light Microtubule-Associated Proteins Muscle Rigidity Protoplasm Radiation Radionuclide Imaging Selenium structural-GP protein, Bos taurus X-Ray Diffraction

Most recents protocols related to «Cyclic Peptides»

Cyclic Peptides Reduce Atherosclerosis

Not available on PMC !

Example 5

Methods.

Experiments were performed in male apoE−/− mice fed an atherogenic diet (D12108, cholate-free AIN-76A semi-purified diet, Research Diets Inc., New Brunswick, NJ) from 4 weeks of age. MPE-298 (300 nmol/kg), MPE-267 (300 nmol/kg) or vehicle (0.9% NaCl), were administered by daily by subcutaneous (s.c.) injections for 8 weeks, from 12 weeks of age, as shown schematically in FIG. 8A.

Results and Discussion.

The results depicted in FIG. 8B show that chronic treatment of the animals with cyclic peptides MPE-267 and MPE-298 reduced aortic lesions by 28% and 42% (p<0.01), respectively, relative to 0.9% NaCl (vehicle), providing evidence that the cyclic peptides exhibit anti-atherosclerotic activity.

US11879021B2. Cyclic peptides and uses thereof (2024-01-23). UNIVERSITÉ DE MONTRÉAL [CA]. Inventors: William D. Lubell [CA], Huy Ong [CA], Jinqiang Zhang [CN], Dilan Mukandila Mulumba [CA], Sylvie Marleau [CA], Ragnhild Gaard Ohm [DK], Ahsanullah [CA], Samy Omri [CA], Ramesh Chingle [US].

Full text: Click here

Patent 2024

Animals Aorta Apolipoproteins E Atherosclerosis Cholate Cyclic Peptides Diet Males Mus Normal Saline

Synthesis of Cyclic Peptide Analogs

Not available on PMC !

Example 2

Tertiary propargylamine bridges were introduced into the peptide by initial incorporation of aza-propargylglycine and ε-N-alkyl-lysine residues into the GHRP-6 peptide sequence, followed by copper-catalyzed macrocyclization using an aldehyde linchpin. The A³-macrocyclization was examined immediately after introduction of the azapropargylglycine residue, as well as after completing the peptide sequence. To seek a diversity-oriented synthesis, two strategies were employed, in which an ε-N-alkyl-lysine residue was introduced respectively at the C-terminal and a central residue of the peptide sequence. With the ε-N-alkyl-lysine residue at the C-terminal, the macrocycle ring-size diversity was varied by azapropargyiglycine position scanning, in which the azapropargylglycyl residue was marched systematically to the N-terminal of the GHRP-6 sequence prior to macrocyclization with formaldehyde. With the ε-N-alkyl-lysine residue centred in the sequence, the influence of various & amino substituents was examined on macrocyclization.

The important step for the effective diversity-oriented synthesis of cyclic azapeptides by A³-macrocyclization was development of solid-phase methods to install the azapropargyiglycine residue and ε-N-alkyl-lysine residue into the peptide sequence prior to the copper-catalyzed macrocyclization using an aldehyde linchpin. The azapropargyiglycine can be inserted by submonomer synthesis of azapeptides on solid phase.^[13]The ε-N-alkylated lysine was prepared in solution and then coupled to the resin-bound peptide; however, solid-phase ε-N-alkylation of lysine was also performed by Mitsunobu chemistry on the corresponding ε-N-o-nitrobenzenesulfonyl (o-NBS) amine.^[20]

As a proof-of-concept of the A³-macrocyclization, cyclic azatripeptide 8 was pursued by placing ε-N-methyl lysine at the peptide C-terminal and inserting aza-propargyiglycine at the i+2 position. Prior to attachment to Rink amide resin, Fmoc-Lys(methyl, o-NBS)—OH 1 was synthesized from Boc-Lys-OH in solution. After Fmoc group removals and elongation with Fmoc-D-Phe-OH using DIC and HOBt, dipeptide 2a was acylated by the active carbazate prepared from benzophenone hydrazone and N,N′-disuccinimidyl carbonate (DSC) to provide semicarbazone 3a.^[14]Propargylation was performed using Cs₂CO₃(300 mol %) and proparyl bromide (600 mol %) to furnish the aza-propargyiglycine 4a in good purity as accessed by LCMS analysis of a cleaved aliquot. After removal of the o-NBS-group with 2-mercaptoethanol and DBU, secondary ε-N-methylamine 5a was ready to test the A³-macrocyclization. Macrocycle 6a was prepared successfully by treating aza-peptide 5a with CuI (20 mol %) and 37% aqueous formaldehyde (600 mol %) in DMSO at rt for 24 h, as verified by LCMS analysis. Elongation of macrocycle 6a to cyclic GHRP-6 analog 8 was accomplished by removal of the semicarbazone with hydroxylamine hydrochloride in pyridine, acylation of the resulting semicarbazide 7a using the symmetric anhydride from treating Fmoc-Ala-OH with DIC, and standard solid-phase peptide synthesis, deprotection and resin cleavage. GHRP-6 macrocycle 8 was isolated in 3.5% overall yield after purification by preparative HPLC. Employing the same strategy, macrocycle 9 was obtained in 2.4% overall yield.

[Figure (not displayed)]

With macrocyclic GHRP-6 analogs 8 and 9 in hand, ring-size scope was investigated by systematically moving the azapropargylglycine residue towards the N-terminal of the sequence. Moreover, the ε-N-alkyl-lysine residue was prepared on solid phase by a method designed to expand the diversity of the ε-amine substituent. After coupling Fmoc-Lys(o-NBS)—OH 10^[19]to RINK amide resin and peptide elongation, semicarbazones 11a-d were synthesized. Chemoselective modification of the ε-N-o-(NBS)amine nitrogen was achieved by employing Mitsunobu chemistry to alkylate the former. Treatment of sulphonamide 11a-d with allyl alcohol, PPh₃, and diisopropyl azodicarboxylate (DIAD) provided selectively ε-N-(allyl)lysinyl peptides 12a-d as verified by LCMS analysis of cleaved aliquots. Subsequently, propargylation of semicarbazone was performed using Cs₂CO₃(300 mol %) and proparyl bromide (600 mol %) to yield aza-propargylglycine peptides 13a-d. A³-Macrocyclization was then performed using the same conditions as discussed above to provide respectively 16-, 19, 21, and 24-membered macrocycles 15a-d as verified by LCMS analysis. After cyclization, semicarbazone removal, semicarbazide acylation, peptide elongation and resin cleavage were performed as described above to afford cyclic GHRP-6 analogs 17 and 18 after purification by preparative HPLC (Table 1). Coupling to semicarbazide macrocycles 16c and 16d was however unsuccessful in the syntheses of the corresponding cyclic GHRP-6 analogs. Steric hindrance inhibited apparently, the coupling to the semicarbazide of the larger ring-sizes. Semicarbazide 16d was however cleaved from resin to give cyclic aza-hexapeptide 19 with a N-terminal semicarbazide after purification by preparative HPLC.

[Figure (not displayed)]
[Figure (not displayed)]
[Figure (not displayed)]

[Figure (not displayed)]
[Figure (not displayed)]

Failure to elongate semicarbazides 16c and 16d after cyclization promoted investigation of a strategy featuring elongation of the complete linear peptide prior to A3-macrocyclization as the penultimate step before simultaneous deprotection and resin cleavage. Semicarbazone 13a was thus treated with hydroxylamine hydrochloride to liberate the semicarbazide 20a, and the linear peptide was elongated as described for its cyclic counterpart above. Aza-hexapeptide 21a was treated with DBU and 2-mercaptoethanol to selectively remove the o-NBS group. Subsequently, aza-hexapeptide 23a was effectively converted to macrocycle 17 using the standard A³-macrocyclization conditions. Resin cleavage gave cyclic azapeptide 17 in about 2-fold higher yield (1.2%) than the earlier approach, involving peptide elongation after cyclization.

Employing the peptide elongation/A³-macrocyclization approach, linear peptides 22b-d were also successfully converted into macrocyclic aza-GHRP-6 analogs 18, 24 and 25. Cyclic azapeptides 24 and 25 were respectively prepared with N-terminal alanine residues to avoid racemization during coupling to the semicarbazide with histidine, and to add an N-terminal basic amine that may favor biological activity.

The diversity of the ε-amine substituent was explored by the synthesis of cyclic azatetrapeptides 30-32 employing different alcohols as electrophiles in the Mitsunobu reaction: methanol, allyl alcohol and isopropyl alcohol. An ε-N-alkylated lysine was inserted in the peptide sequence to replace the tryptophan residue and an azapropargylglycine was placed at the i+3 position to replace the histidine residue in the GHRP-6 sequence. Cyclic analog 33 was synthesized with an additional alanine in the N-terminal for comparison with analog 31 to study the importance of the N-terminal basic amine.

[Figure (not displayed)]

Cyclic azapeptide GHRP-6 analogs were synthesized by the A³-macrocyclization method in yields and purities suitable for biological evaluation (Table 1).

TABLE 1

Yields and purity of the cyclic azapeptide GHRP-6 analogs

CyclicSyntheticIsolated

AnalogApproachYield (%)Purity^[a]HRMS

8I3.5 99%809.4201(809.4206)

9I2.4 99%924.4627(924.4628)

17I and (II)0.5(1.5)99%835.4376(835.4362)

18I and (II)0.4(1.1)99%950.4787(950.4784)

19I0.5 94%884.4549(884.4566)

24II0.9 99%769.4140(769.4144)

25II1.1 97%955.4942(955.4937)

26II2.0%99%997.5031(997.5043)

27II1.6%99%926.4658(926.4671)

31I1.5 96%826.4718(826.4723)

32I1.2 97%828.4875(828.4879)

33II0.9 94%897.5092(897.5094)

34II2.5%98%939.5186(939.5199)

35II1.4%96%868.4804(868.4828)

^[a]Determined by LCMS analysis as described above.

Synthesis of Cyclic Analogs MPE-110, MPE-111, MPE-074 and MPE-048

Solution-Phase Chemistry

Ornithine Building Block Synthesis

[Figure (not displayed)]

Fmoc-Orn(o-NBS)—OH (RGO1):

Fmoc-Orn(Boc)-OH (2.02 g, 4.44 mmol) was dissolved in CH₂Cl₂(30 mL) treated with TFA (20 mL) stirred at room temperature for 3 hours, and the volatiles were removed by rotary evaporation. The resulting yellow oil was co-evaporated with toluene to give a residue that was dissolved in THF (40 mL) and water (40 mL) and treated with iPr₂NEt (7.70 mL, 44.2 mmol) and o-NBSCl (1.13 g, 5.08 mmol) in one portion. The reaction was stirred at room temperature for 3 hours, diluted with EtOAc (100 mL) and sequentially washed with aqueous HCl (1 M, 100 mL×3), water (100 mL) and brine (100 mL). The organic layer was dried over MgSO₄and the volatiles were removed by rotary evaporation to give sulfonamide RGO1 (2.4 g, quant.) as a light yellow solid. The amino acid was used without further purification.

¹H NMR (300 MHz, DMSO) δ 8.10 (t, J=5.6 Hz, 1H), 8.03-7.92 (m, 2H), 7.92-7.81 (m, 4H), 7.72 (d, J=7.4 Hz, 2H), 7.61 (d, J=8.0 Hz, 1H), 7.41 (t, J=7.2 Hz, 2H), 7.32 (t, J=7.1 Hz, 2H), 4.34-4.16 (m, 3H), 3.89 (td, J=8.7, 4.6 Hz, 1H), 2.90 (q, J=6.3 Hz, 2H), 1.73 (s, 1H), 1.65-1.43 (m, 3H). ¹³C NMR (75 MHz, DMSO) δ 173.7, 156.1, 147.8, 143.8, 140.7, 134.0, 132.7, 132.6, 129.4, 127.7, 127.1, 125.3, 124.4, 120.1, 65.6, 53.5, 46.7, 42.3, 27.9, 26.0. LCMS (10-90% MeOH containing 0.1% formic acid over 10 min) R_t=11.04 min. ESI-MS m/z calcd for C₂₆H₂₆N₃O₈S⁺ [M+H]⁺ 540.1, found 540.1. Melting point: 108-110° C.

Solid-Phase Chemistry

Fmoc-based peptide synthesis was performed using standard conditions (W. D. Lubell, J. W. Blankenship, G. Fridkin, and R. Kaul (2005) “Peptides.” Science of Synthesis 21.11, Chemistry of Amides. Thieme, Stuttgart, 713-809) on an automated shaker using polystyrene Rink amide resin. The loading was calculated from the UV absorbance for Fmoc-deprotection after the coupling of the first amino acid. Couplings of amino acids (3 equiv.) were performed in DMF using DIC (3 equiv.) and HOBt (3 equiv.) for 3-6 hours. Fmoc-deprotections were performed by treating the resin with 20% piperidine in DMF for 30 min. The resin was washed after each coupling and deprotection step sequentially with DMF (×3), MeOH (×3) THF (×3) and CH₂Cl₂(×3).

Lysine as AA1

[Figure (not displayed)]

Fmoc-Lys(o-NBS)-Rink Amide Resin RGO7:

On Rink amide resin (3.00 g) in a syringe fitted with a Teflon™ filter, Fmoc removal was performed by treating the resin with a solution of 20% piperidine in DMF over 30 min. The resin was filtered and washed sequentially with DMF (×3), MeOH (×3) and CH₂Cl₂(×3). Fmoc-Lys(o-NBS)—OH (1.62 g, 2.93 mmol) was dissolved in DMF (20 mL) and treated with DIC (0.7 mL, 4.52 mmol) and HOBt (611 mg, 4.52 mmol), stirred for 3 min. and added to the syringe containing the resin. The mixture was shaken for 14 hours. The resin was then filtered and sequentially washed with DMF (×3), MeOH (×3) and CH₂Cl₂(×3). The resin was dried and the loading was measured at 0.345 mmol/g resin.

[Figure (not displayed)]

Fmoc-Lys(o-NBS, Allyl)-Rink Amide Resin RGO8:

Vacuum dried Fmoc-Lys(o-NBS)-resin RGO7 (0.441 mmol) was placed in a syringe fitted with a Teflon™ filter, suspended in THF (dry, 5 mL) and treated sequentially with solutions of allyl alcohol (206 μL, 3.03 mmol) in THF (dry, 1 mL), PPh₃(397 mg, 1.51 mmol) in THF (dry, 1 mL), and DIAD (298 μL, 1.51 mmol) in THF (dry, 1 mL). The mixture in the syringe was shaken for 90 min. The resin was filtered and sequentially washed with DMF (×3), MeOH (×3), THF (×3) and CH₂Cl₂(×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete allylation: LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=8.65 min. ESI-MS m/z calcd for C₃₀H₃₃N₄O₇S⁺ [M+H]⁺593.2, found 593.2.

[Figure (not displayed)]

Boc-Ala-D-Pra-Ala-Trp(Boc)-D-Phe-Lys(o-NBS, Allyl)-Rink Amide Resin RGO99:

LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=5.73 min. ESI-MS m/z calcd for C₄₆H₅₇N₁₀O₁₀S⁺ [M−2Boc+H]⁺941.4, found 941.4.

[Figure (not displayed)]

Boc-Ala-L-Pra-Ala-Trp(Boc)-D-Phe-Lys(o-NBS, Allyl)-Rink Amide Resin RGO100:

LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=5.77 min. ESI-MS m/z calcd for C₄₆H₅₇N₁₀O₁₀S⁺ [M−2Boc+H]⁺941.4, found 941.4.

[Figure (not displayed)]

Boc-Ala-D-Pra-D-Trp(Boc)-Ala-Trp-D-Phe-Lys(o-NBS, Allyl)-Rink Amide Resin RGO65:

LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=6.48 min. ESI-MS m/z calcd for C₅₇H₆₇N₁₂O₁₁S⁺ [M−3Boc+H]⁺1127.5, found 1127.5.

[Figure (not displayed)]

Boc-Ala-L-Pra-D-Trp(Boc)-Ala-Trp-D-Phe-Lys(o-NBS, Allyl)-Rink Amide Resin RGO66:

LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=6.66 min. ESI-MS m/z calcd for C₅₇H₆₇N₁₂O₁₁S⁺ [M−3Boc+H]⁺1127.5, found 1127.5.

[Figure (not displayed)]

Boc-Ala-D-Pra-Ala-Trp(Boc)-D-Phe-Lys(Allyl)-Rink Amide Resin RGO104:

o-NBS-protected hexapeptide RGO99 (˜600 mg, 0.156 mmol) in a syringe fitted with a Teflon™ filter was swollen in DMF (5 mL) and treated with DBU (210 μL, 1.40 mmol) and 2-mercaptoethanol (50 μL, 0.71 mmol). The mixture in the syringe was shaken for 1 h. The resin was filtered and sequentially washed with DMF (×3), MeOH (×3), THF (×3) and CH₂Cl₂(×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete o-NBS-removal: LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=1.50 min. ESI-MS m/z calcd for C₄₀H₅₄N₉O₆⁺ [M−2Boc+H]⁺756.4, found 756.4.

[Figure (not displayed)]

Boc-Ala-L-Pra-Ala-Trp(Boc)-D-Phe-Lys(Allyl)-Rink Amide Resin RGO105:

o-NBS-protected hexapeptide RGO100 (˜600 mg, 0.14 mmol) in a syringe fitted with a Teflon™ filter was swollen in DMF (5 mL) and treated with DBU (210 μL, 1.40 mmol) and 2-mercaptoethanol (50 μL, 0.71 mmol). The mixture in the syringe was shaken for 1 h. The resin was filtered and sequentially washed with DMF (×3), MeOH (×3), THF (×3) and CH₂Cl₂(×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete o-NBS-removal: LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=1.51 min. ESI-MS m/z calcd for C₄₀H₅₄N₉O₆⁺ [M−2Boc+H]⁺756.4, found 756.4.

[Figure (not displayed)]

Boc-Ala-D-Pra-D-Trp(Boc)-Ala-Trp(Boc)-D-Phe-Lys(allyl)-Rink Amide Resin RGO69:

o-NBS-protected heptapeptide RGO65 (˜300 mg, 0.10 mmol) in a syringe fitted with a Teflon™ filter was swollen in DMF (6 mL) and treated with DBU (150 μL, 1.00 mmol) and 2-mercaptoethanol (35 μL, 0.50 mmol). The mixture in the syringe was shaken for 1 h. The resin was filtered and sequentially washed with DMF (×3), MeOH (×3), THF (×3) and CH₂Cl₂(×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete o-NBS-removal: LCMS (20-80% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=4.79 min. ESI-MS m/z calcd for C₅₁H₆₄N₁₁O₇⁺ [M−3Boc+H]⁺942.5, found 942.5.

[Figure (not displayed)]

Boc-Ala-L-Pra-D-Trp(Boc)-Ala-Trp(Boc)-D-Phe-Lys(Allyl)-Rink Amide Resin RGO70:

o-NBS-protected heptapeptide RGO66 (˜300 mg, 0.09 mmol) in a syringe fitted with a Teflon™ filter was swollen in DMF (6 mL) and treated with DBU (130 μL, 0.87 mmol) and 2-mercaptoethanol (30 μL, 0.43 mmol). The mixture in the syringe was shaken for 1 h. The resin was filtered and sequentially washed with DMF (×3), MeOH (×3), THF (×3) and CH₂Cl₂(×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete o-NBS-removal: LCMS (20-80% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=5.05 min. ESI-MS m/z calcd for C₅₁H₆₄N₁₁O₇⁺ [M−3Boc+H]⁺942.5, found 942.5.

[Figure (not displayed)]

Cyclic Peptide MPE-110:

Hexapeptide resin RGO104 (˜600 mg, 0.156 mmol) was swollen in DMSO (6 mL) for 30 min in a syringe tube equipped with Teflon™ filter, and stopper, treated with CuI (5.0 mg, 0.03 mmol) and aqueous formaldehyde (70 μL, 0.94 mmol, 37% in H₂O), shaken on an automated shaker for 30 h, and filtered. After filtration, the resin was washed sequentially with AcOH/H₂O/DMF (5:15:80, v/v/v, ×3), DMF (×3), THF (×3), MeOH (×3), and DCM (×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete conversion, and a peak with molecular ion consistent with cyclic hexapeptide MPE-110 was observed: MS m/z calcd for Ca₄₁H₅₄N₉O₆⁺ [M+H]⁺ 768.4, found 768.4.

Resin-bound cyclic peptide MPE-110 was deprotected and cleaved from the support using a freshly made solution of TFA/H₂O/TES (95:2.5:2.5, v/v/v, 5 mL) at rt for 2 h. The resin was filtered and rinsed with TFA (5 mL). The filtrate and rinses were concentrated until a crude oil persisted, from which a precipitate was obtained by addition of cold ether (10 mL). After centrifugation (1200 rpm for 10 min), the supernatant was removed and the crude peptide precipitate was taken up in aqueous MeOH (10% v/v) and freeze-dried prior to purification. The resulting light brown fluffy material was purified by preparative HPLC to give cyclic pentapeptide MPE-110 (2.0 mg, 2%) as white fluffy material.

LCMS analysis of cyclic peptide MPE-110 was performed using a linear gradient of a) 10-90% of MeOH containing 0.1% formic acid in H₂O (0.1% formic acid) over 10 min, then at 10% MeOH (0.1% formic acid) for 5 min, R_t=4.24 min; b) 10-90% MeCN containing 0.1% formic acid in H₂O containing 0.1% formic acid over 10 min, then at 10% MeCN (0.1% formic acid) for 5 min, R_t=1.70 min; HRMS m/z. calcd for C₄₁H₅₄N₉O₆⁺ [M+H]⁺768.4192, found 768.4176.

[Figure (not displayed)]

Cyclic Peptide MPE-111:

Hexapeptide resin RGO105 (˜600 mg, 0.14 mmol) was swollen in DMSO (6 mL) for 30 min in a syringe tube equipped with Teflon™ filter, and stopper, treated with CuI (5.0 mg, 0.03 mmol) and aqueous formaldehyde (60 μL, 0.84 mmol, 37% in H₂O), shaken on an automated shaker for 30 h, and filtered. After filtration, the resin was washed sequentially with AcOH/H₂O/DMF (5:15:80, v/v/v, ×3), DMF (×3), THF (×3), MeOH (×3), and DCM (×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete conversion, and a peak with molecular ion consistent with cyclic hexapeptide MPE-111 was observed: MS m/z. calcd for C₄₁H₅₄N₉O₆⁺ [M+H]⁺ 768.4, found 768.4.

Resin-bound cyclic peptide MPE-111 was deprotected and cleaved from the support using a freshly made solution of TFA/H₂O/TES (95:2.5:2.5, v/v/v, 5 mL) at rt for 2 h. The resin was filtered and rinsed with TFA (5 mL). The filtrate and rinses were concentrated until a crude oil persisted, from which a precipitate was obtained by addition of cold ether (10 mL). After centrifugation (1200 rpm for 10 min), the supernatant was removed and the crude peptide precipitate was taken up in aqueous MeOH (10% v/v) and freeze-dried prior to purification. The resulting light brown fluffy material was purified by preparative HPLC to give cyclic hexapeptide MPE-111 (2.9 mg, 3%) as white fluffy material.

LCMS analysis of cyclic peptide MPE-111 was performed using a linear gradient of a) 10-90% of MeOH containing 0.1% formic acid in H₂O (0.1% formic acid) over 10 min, then at 10% MeOH (0.1% formic acid) for 5 min, R_t=4.50 min; b) 10-90% MeCN containing 0.1% formic acid in H₂O containing 0.1% formic acid over 10 min, then at 10% MeCN (0.1% formic acid) for 5 min, R_t=2.03 min; HRMS m/z. calcd for C₄₁H₅₄N₉O₆⁺ [M+H]⁺ 768.4192, found 768.4172.

[Figure (not displayed)]

Cyclic Peptide MPE-074:

Heptapeptide resin RGO69 (˜300 mg, 0.10 mmol) was swollen in DMSO (5 mL) for 30 min in a syringe tube equipped with Teflon™ filter, and stopper, treated with CuI (4.0 mg, 0.02 mmol) and aqueous formaldehyde (50 μL, 0.69 mmol, 37% in H₂O), shaken on an automated shaker for 29 h, and filtered. After filtration, the resin was washed sequentially with AcOH/H₂O/DMF (5:15:80, v/v/v, ×3), DMF (×3), THF (×3), MeOH (×3), and DCM (×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete conversion, and a peak with molecular ion consistent with cyclic heptapeptide MPE-074 was observed: MS m/z calcd for C₅₂H₆₃N₁₁NaO₇⁺ [M+Na]⁺976.5, found 976.4.

Resin-bound cyclic peptide MPE-074 was deprotected and cleaved from the support using a freshly made solution of TFA/H₂O/TES (95:2.5:2.5, v/v/v, 5 mL) at rt for 2 h. The resin was filtered and rinsed with TFA (5 mL). The filtrate and rinses were concentrated until a crude oil persisted, from which a precipitate was obtained by addition of cold ether (10 mL). After centrifugation (1200 rpm for 10 min), the supernatant was removed and the crude peptide precipitate was taken up in aqueous MeOH (10% v/v) and freeze-dried prior to purification. The resulting light brown fluffy material was purified by preparative HPLC to give cyclic heptapeptide MPE-074 (0.7 mg, 1%) as white fluffy material.

LCMS analysis of cyclic peptide MPE-074 was performed using a linear gradient of a) 10-90% of MeOH containing 0.1% formic acid in H₂O (0.1% formic acid) over 10 min, then at 10% MeOH (0.1% formic acid) for 5 min, R_t=1.72 min; b) 10-90% MeCN containing 0.1% formic acid in H₂O containing 0.1% formic acid over 10 min, then at 10% MeCN (0.1% formic acid) for 5 min, R_t=4.24 min; HRMS m/z calcd for C₅₂H₆₃N₁₁NaO₇⁺ [M+Na]⁺976.4804, found 976.4817.

[Figure (not displayed)]

Cyclic Peptide MPE-075:

Heptapeptide resin RGO69 (˜300 mg, 0.09 mmol) was swollen in DMSO (5 mL) for 30 min in a syringe tube equipped with Teflon™ filter, and stopper, treated with CuI (3.0 mg, 0.02 mmol) and aqueous formaldehyde (50 μL, 0.69 mmol, 37% in H₂O), shaken on an automated shaker for 29 h, and filtered. After filtration, the resin was washed sequentially with AcOH/H₂O/DMF (5:15:80, v/v/v, ×3), DMF (×3), THF (×3), MeOH (×3), and DCM (×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete conversion, and a peak with molecular ion consistent with cyclic heptapeptide MPE-075 was observed: MS m/z calcd for C₅₂H₆₄N₁₁O₇⁺ [M+H]⁺ 954.5, found 954.5.

Resin-bound cyclic peptide MPE-075 was deprotected and cleaved from the support using a freshly made solution of TFA/H₂O/TES (95:2.5:2.5, v/v/v, 5 mL) at rt for 2 h. The resin was filtered and rinsed with TFA (5 mL). The filtrate and rinses were concentrated until a crude oil persisted, from which a precipitate was obtained by addition of cold ether (10 mL). After centrifugation (1200 rpm for 10 min), the supernatant was removed and the crude peptide precipitate was taken up in aqueous MeOH (10% v/v) and freeze-dried prior to purification. The resulting light brown fluffy material was purified by preparative HPLC to give cyclic heptapeptide MPE-075 (1.5 mg, 2%) as a white fluffy material.

LCMS analysis of cyclic peptide MPE-075 was performed using a linear gradient of a) 10-90% of MeOH containing 0.1% formic acid in H₂O (0.1% formic acid) over 10 min, then at 10% MeOH (0.1% formic acid) for 5 min, R_t=1.89 min; b) 10-90% MeCN containing 0.1% formic acid in H₂O containing 0.1% formic acid over 10 min, then at 10% MeCN (0.1% formic acid) for 5 min, R_t=4.47 min; HRMS m/z calcd for C₅₂H₆₄N₁₁O₇⁺ [M+H]⁺ 954.4985, found 954.4973.

Ornithine as AA1

[Figure (not displayed)]

Fmoc-Orn(o-NBS)-Rink Amide Resin RGO3:

Rink amide resin (2.51 g) was placed in a syringe fitted with a Teflon™ filter. The Fmoc group was removed by treating the resin with a solution of 20% piperidine in DMF over 30 min. The resin was filtered and washed sequentially with DMF (×3), MeOH (×3) and CH₂Cl₂(×3). Fmoc-Orn(o-NBS)—OH (1.33 g, 2.46 mmol) was dissolved in DMF (20 mL) and treated with DIC (0.57 mL, 3.68 mmol) and HOBt (494 mg, 3.66 mmol) and stirred for 3 min, before being transferred to the syringe containing the swollen resin, and the mixture was shaken for 14 hours. The resin was filtered and washed sequentially with DMF (×3), MeOH (×3) and CH₂Cl₂(×3). The resin was dried and the loading was measured to 0.187 mmol/g resin.

[Figure (not displayed)]

Fmoc-Orn(o-NBS, Allyl)-Rink Amide Resin RGO4:

Vacuum dried Fmoc-Orn(o-NBS)-resin (0.362 mmol) was placed in a syringe fitted with a Teflon™ filter, suspended in THF (dry, 20 mL) and treated sequentially with solutions of allyl alcohol (250 μL, 3.68 mmol) in THF (dry, 1 mL), PPh₃(482 mg, 1.84 mmol) in THF (dry, 2 mL) and DIAD (360 μL, 1.83 mmol) in THF (dry, 1 mL). The resin mixture in the syringe was shaken for 90 min. The resin was filtered and washed sequentially with DMF (×3), MeOH (×3), THF (×3) and CH₂Cl₂(×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete allylation: LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=8.47 min. ESI-MS m/z calcd for C₂₉H₃₁N₄O₇S⁺ [M+H]⁺579.2, found 579.2.

[Figure (not displayed)]

Fmoc-D-Trp(Boc)-Ala-Trp(Boc)-D-Phe-Orn(o-NBS, Allyl)-Rink Amide Resin RGO22:

LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=6.13 min. ESI-MS m/z calcd for C₄₈H₅₅N₁₀O₉S⁺ [M-Fmoc-2Boc+H]⁺947.4, found 947.3.

[Figure (not displayed)]

Fmoc-azaPra-D-Trp(Boc)-Ala-Trp(Boc)-D-Phe-Orn(o-NBS, Allyl)-Rink Amide Resin RGO79:

N′-Propargyl-fluorenylmethylcarbazate (248 mg, 0.849 mmol, prepared by alkylation of fluorenylmethylcarbazate with propargyibromide as —N(R¹⁰)— described below) was dissolved in CH₂Cl₂(dry, 40 mL) under argon atmosphere. The solution was cooled to 0° C., treated with a 20% solution of phosgene in toluene (1 mL, 1.87 mmol), warmed to rt, stirred 50 min, and the volatiles were removed by rotary evaporation. The residue was re-dissolved in CH₂Cl₂(10 mL) and the volatiles were once again removed by rotary evaporation. The resulting white solid was dissolved in CH₂Cl₂(dry, 7 mL) and added to the Fmoc-deprotected pentapeptide RGO22 in a syringe fitted with a Teflon™ filter. The mixture in the syringe was shaken for 28 h. The resin was filtered and washed sequentially with DMF (×3), MeOH (×3), THF (×3) and CH₂Cl₂(×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete coupling: LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=8.26 min. ESI-MS m/z calcd for C₅₂H₅₉N₁₂O₁₀S⁺ [M-Fmoc-2Boc+H]⁺1043.4, found 1043.3.

[Figure (not displayed)]

Boc-Ala-azaPra-D-Trp(Boc)-Ala-Trp(Boc)-D-Phe-Orn(o-NBS, Allyl)-Rink Amide RGO29:

Coupling onto the semicarbazide RGO79 was performed by using amino acid symmetric anhydrides that were generated in situ (J. Zhang, C. Proulx, A. Tomberg, W. D. Lubell, Org. Lett. 2013, 16, 298-301). The procedure was repeated twice on semicarbazide RGO79. Examination by LCMS of a cleaved resin sample (5 mg) showed complete coupling: LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=6.39 min. ESI-MS m/z calcd for C₅₅H₆₄N₁₃O₁₁S⁺ [M−3Boc+H]⁺1114.5, found 1114.4.

[Figure (not displayed)]

Boc-Ala-azaPra-D-Trp(Boc)-Ala-Trp(Boc)-D-Phe-Orn(Allyl)-Rink Amide RGO30:

o-NBS-protected hetapeptide RGO29 (˜1 g, 0.2 mmol) in a syringe fitted with a Teflon™ filter was swollen in DMF (6 mL) and DBU (300 μL, 2.01 mmol) and treated with 2-mercaptoethanol (70 μL, 1.00 mmol). The mixture in the syringe was shaken for 1 h. The resin was filtered and washed sequentially with DMF (×3), MeOH (×3), THF (×3) and CH₂Cl₂(×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete o-NBS-removal: LCMS (30-95% MeOH containing 0.1% formic acid in water containing 0.1% formic acid over 10 min) R_t=4.49 min. ESI-MS m/z calcd for C₄₉H₆₁N₁₂O₇⁺ [M−3Boc+2Na]²⁺487.2, found 487.3.

[Figure (not displayed)]

Cyclic Azapeptide MPE-048:

Azaheptapeptide resin RGO30 (˜1 g, 0.2 mmol) was swollen in DMSO (8 mL) for 30 min in a syringe tube equipped with Teflon™ filter, and stopper, treated with CuI (7.0 mg, 0.04 mmol) and aqueous formaldehyde (90 μL, 1.2 mmol, 37% in H₂O), shaken on an automated shaker for 31 h, and filtered. After filtration, the resin was washed sequentially with AcOH/H₂O/DMF (5:15:80, v/v/v, ×3), DMF (×3), THF (×3), MeOH (×3), and DCM (×3). Examination by LCMS of a cleaved resin sample (5 mg) showed complete conversion, and a peak with molecular ion consistent with cyclic azaheptapeptide MPE-048 was observed: MS m/z calcd for C₅₀H₆₁N₁₂O₇⁺[M+H]⁺941.5, found 941.4.

Resin-bound cyclic azapeptide MPE-048 was deprotected and cleaved from the support using a freshly made solution of TFA/H₂O/TES (95:2.5:2.5, v/v/v, 5 mL) at rt for 2 h. The resin was filtered and rinsed with TFA (5 mL). The filtrate and rinses were concentrated until a crude oil persisted, from which a precipitate was obtained by addition of cold ether (10 mL). After centrifugation (1200 rpm for 10 min), the supernatant was removed and the crude peptide precipitate was taken up in aqueous MeOH (10% v/v) and freeze-dried prior to purification. The resulting light brown fluffy material was purified by preparative HPLC to give cyclic azaheptapeptide MPE-048 (1.3 mg, 1%) as white fluffy material.

LCMS analysis of cyclic peptide MPE-048 was performed using a linear gradient of a) 10-90% of MeOH containing 0.1% formic acid in H₂O (0.1% formic acid) over 10 min, then at 10% MeOH (0.1% formic acid) for 5 min, R_t=1.80 min; b10-90% MeCN containing 0.1% formic acid in H₂O containing 0.1% formic acid over 10 min, then at 10% MeCN (0.1% formic acid) for 5 min, R_t=4.30 min; HRMS m/z calcd for C₅₀H₆₀N₁₂O₇Na⁺ [M+Na]⁺963.4600, found 963.4573.

Synthesis of Cyclic Analogs MPE-189, MPE-201, MPE-202, and MPE-203

Synthesis of Carbazates 2 and 3

[Figure (not displayed)]

To a well-stirred solution of fluorenylmethyl carbazate (1, 1 eq., 2.8 g, 11 mmol, prepared according to reference 1) and DIEA (2 eq., 2.85 g, 3.64 mL, 22 mmol) in dry DMF (280 mL) at 0° C., a solution of 3-bromopropyne (0.9 eq., 1.47 g, 1.07 mL, 9.91 mmol, 80 wt. % in toluene) in dry DMF (10 mL) was added drop-wise by cannula over 30 min. The cooling bath was removed. The reaction mixture was allowed to warm to room temperature and stirred for 16 h. The volatiles were evaporated. The residue was partitioned between EtOAc and brine. The aqueous layer was separated and extracted with EtOAc. The combined organic layer was dried over Na₂SO₄, filtered, and evaporated. The residue was purified by silica gel chromatography eluting with 40% EtOAc in hexane as solvent system to obtain N′-propargyl-fluorenylmethylcarbazate 3 (1.8 g, 62%), as white solid: R_f0.42 (60% EtOAc); mp 148-149° C.; ¹H NMR (500 MHz, DMSO-d₆) δ 8.82 (s, 1H), 7.89 (d, J=7.5 Hz, 2H), 7.70 (d, J=7.4 Hz, 2H), 7.50-7.43 (m, 2H), 7.37-7.28 (m, 2H), 4.89 (q, J=4.6 Hz, 1H), 4.29 (d, J=6.9 Hz, 2H), 4.22 (t, J=6.1 Hz, 1H), 3.48 (s, 2H), 3.09 (t, J=2.3 Hz, 1H); ¹³C NMR (125 MHz, DMSO-d₆) δ 156.7, 143.8, 140.7, 127.7 (2C), 127.1 (2C), 125.3 (2C), 120.1 (2C), 81.2, 74.2, 65.6 (2C), 46.6, 39.6 (2C). IR (neat) vmax/cm-1 3304, 3290, 2947, 1699, 1561, 1489, 1448, 1265, 1159, 1021; HRMS m/z calculated for C₁₈H₁₇N₂O₂[M+H]⁺293.1285; found 293.1275.

Full text: Click here

Patent 2024

1-hydroxybenzotriazole 1H NMR 2-Mercaptoethanol 5A peptide Acylation Alanine Alcohols Aldehydes Alkylation allyl alcohol Amides Amines Amino Acids Anabolism Anhydrides Argon Atmosphere Bath benzophenone Biopharmaceuticals brine Bromides Cannula carbamylhydrazine carbazate Carbon-13 Magnetic Resonance Spectroscopy Carbonates Cardiac Arrest Centrifugation Chromatography Cold Temperature Copper Cyclic Peptides Cyclization Cytokinesis Dipeptides Ethers Filtration Formaldehyde formic acid Freezing Gel Chromatography growth hormone releasing hexapeptide H 1285 Hexanes High-Performance Liquid Chromatographies Histidine Hydrazones Hydroxylamine Hydroxylamine Hydrochloride Isopropyl Alcohol Light Lincomycin Methanol methylamine N,N-diisopropylethylamine N-propargyl Nitrogen Ornithine Peptide Biosynthesis Peptides Petroleum Phosgene piperidine polypeptide C Polystyrenes propargylamine propargylglycine pyridine pyridine hydrochloride Resins, Plant Rink amide resin Semicarbazides Semicarbazones Silica Gel Silicon Dioxide Solvents Sulfate, Magnesium Sulfonamides Sulfoxide, Dimethyl Syringes Teflon tert-butoxycarbonylalanine Toluene Training Programs Tryptophan Vacuum

Rosetta Cyclic Peptide Energy Landscape

Cited 1 time

Energy landscape calculations for cyclic peptides were done as previously described using Rosetta cycpep_predict application (18 (link)–20 (link)). Large-scale conformational sampling during these calculations was conducted using the BOINC Rosetta@Home platform. Energy for each sampled conformation was calculated using the Rosetta REF2015 energy function(29 (link), 30 (link)). The folding propensity was evaluated based on the energy gap between the design conformation and alternative conformations, and by calculating P_near, a Rosetta metric that looks at the quality of energy ‘funnel’. P_near value of 1 denotes energy landscapes with a funnel that converges to the designed model as its single low-energy minima; 0 denotes energy landscapes with one or more alternative conformations as energy minima that are different from the designed conformation. See ref (20 (link)) for details of the P_near metric.

Rettie S.A., Campbell K.V., Bera A.K., Kang A., Kozlov S., De La Cruz J., Adebomi V., Zhou G., DiMaio F., Ovchinnikov S, & Bhardwaj G. (2023). Cyclic peptide structure prediction and design using AlphaFold. bioRxiv.

Publication Preprint 2023

Cyclic Peptides

Osteoblast Attachment and OPN Expression on Titanium Surfaces

Osteoblasts were inoculated onto the Ti and Ti-gel/n (CAT) surfaces at a density of 1 × 10⁴ cells per well according to the previous method. After 2 days of incubation, the α-MEM medium was replaced with an osteogenic induction medium. The medium was changed daily until day 7, when immunofluorescence staining for OPN was performed. Briefly, cells were fixed in 4% tissue fixative for 15 min, rinsed three times with 1 x PBS solution, followed by further treatment of the samples with 0.3%–0.5% Triton X-100, immediately followed by the addition of 10% goat serum for 1 h at room temperature to block non-specific binding. After adding primary antibodies to the samples overnight at 4°C, secondary antibodies Alexa Fluor 647 goat anti-mouse IgG (Abcam) were added and incubated at room temperature for 2 h. The cytoskeleton and nuclei were then treated with rhodamine-labeled ghost pencil cyclic peptide and DAPI. Finally, the OPN fluorescence images of the surfaces of different materials were observed by a fluorescence confocal microscope. (Nikon).

Zhang T., Qin X., Gao Y., Kong D., Jiang Y., Cui X., Guo M., Chen J., Chang F., Zhang M., Li J, & Yin P. (2023). Functional chitosan gel coating enhances antimicrobial properties and osteogenesis of titanium alloy under persistent chronic inflammation. Frontiers in Bioengineering and Biotechnology, 11, 1118487.

Full text: Click here

Publication 2023

Alexa Fluor 647 anti-IgG Antibodies Cardiac Arrest Cell Nucleus Cells Cyclic Peptides Cytoskeleton DAPI Fixatives Fluorescence Fluorescent Antibody Technique Goat Microscopy, Fluorescence Mus Osteoblasts Osteogenesis Red Cell Ghost Rhodamine Serum Specimen Handling Tissues Triton X-100

Radiolabeled DX600 Peptide Synthesis and Evaluation

⁶⁸Ga-cyc-DX600 was prepared in house with DOTA-modified cyclic DX600 peptide (cyc-DX600-DOTA, DX600 peptide with condensed disulfide bond of cysteine) as the precursor following the reported protocol [11 (link), 12 ]. In detail, the newly eluted ⁶⁸Ga³⁺ in 4 mL 0.05 M HCl was mixed with precursor in 1 mL 0.25 M NH₄Ac, heated to 100 ℃ and maintained for 10 min. Radiochemical purity (RCP) was measured with HPLC system (1260 Infinity, Agilent Technologies) equipped with a radioactive detector (Flow-count, Eckert & Ziegler) to determine the labeling rate, as well as the stability in PBS and 5% fetal bovine serum for one hour incubation at room temperature.
¹⁸F-FDG was purchased from Atom Kexing Radiopharmaceuticals Ltd with strict quality control. Radiopharmaceuticals with labeling rate higher than 95% were used in imaging research immediately, and the specific radioactivity of ⁶⁸Ga-cyc-DX600 was controlled as about 3.7 MBq/µg in the imaging research on various tumor-bearing mice models or patients.
To verify the maintain of ACE2 targeting ability on cellular level, ¹²⁵I-labeled DX600 and ¹²⁵I-labeled cyc-DX600-DOTA were synthesized via Iodogen-catalyzed iodization with I-125. Cellular binding was performed on 1 × 10⁵ HEK-293T/hACE2 cells that expressed humanized ACE2 protein. After the co-culture with 1 nmol radiopharmaceuticals for 30 min or 1 h, the excess radiopharmaceuticals were washed away by cold PBS for three times, and the bound peptide or precursor was quantified with a gamma-detector.

Ren F., Jiang H., Shi L., Zhang L., Li X., Lu Q, & Li Q. (2023). 68Ga-cyc-DX600 PET/CT in ACE2-targeted tumor imaging. European Journal of Nuclear Medicine and Molecular Imaging, 50(7), 2056-2067.

Publication 2023

ACE2 protein, human Angiotensin Converting Enzyme 2 Cells Coculture Techniques Cold Temperature Cyclic Peptides Cystine DX600 peptide F18, Fluorodeoxyglucose Fetal Bovine Serum Gamma Rays HEK293 Cells High-Performance Liquid Chromatographies Iodine-125 Iodo-Gen Mus Neoplasms Patients Peptides Radioactivity Radiopharmaceuticals tetraxetan

Top products related to «Cyclic Peptides»

J 815 cd spectrometer by Jasco

Sourced in Japan, United States, United Kingdom, Germany, Canada, Brazil

The J-815 CD spectrometer is a laboratory instrument designed to measure the circular dichroism (CD) spectrum of samples. It is capable of analyzing the interactions between light and optically active molecules, providing information about the structural and conformational properties of the sample. The J-815 CD spectrometer operates within a specified wavelength range and can be used to study a variety of biomolecules, such as proteins, nucleic acids, and small organic compounds.

Milli q water by Merck Group

Sourced in United States, Germany, France, Australia, United Kingdom, India, Japan, Italy, Spain, Switzerland, China, Belgium, Netherlands, Canada, Czechia, Austria, Morocco, Brazil, Denmark, Poland

Milli-Q water is a high-purity water purification system produced by Merck Group. It uses a combination of filtration, ion exchange, and other purification techniques to remove impurities and produce water with low levels of dissolved solids, organics, and microorganisms.

J 815 spectropolarimeter by Jasco

Sourced in Japan, United States, Germany, United Kingdom, Italy, Canada

The J-815 spectropolarimeter is a laboratory instrument used for the measurement of circular dichroism (CD) spectra. It is designed to analyze the interaction between polarized light and chiral molecules or structures. The J-815 spectropolarimeter provides accurate and reliable data on the secondary structure and conformational changes of biological macromolecules such as proteins, nucleic acids, and other chiral compounds.

J 810 spectropolarimeter by Jasco

Sourced in Japan, United States, Italy, Germany, United Kingdom, Canada, France

The J-810 spectropolarimeter is a laboratory instrument designed for the measurement of circular dichroism (CD) and other optical rotatory properties of samples. It provides accurate and reliable data on the secondary structure and conformational changes of biological macromolecules such as proteins, nucleic acids, and other chiral compounds.

Voyager workstation by Thermo Fisher Scientific

The Voyager workstation is a lab equipment product designed for sample preparation and processing. It features a compact and modular design to accommodate various laboratory workflows.

415 autosampler by AB Sciex

Sourced in United States

The 415 autosampler is a precision liquid handling instrument designed for automated sample injection in analytical workflows. It features a compact design and offers reliable and consistent sample handling capabilities for a wide range of applications.

Fetal bovine serum (fbs) by Thermo Fisher Scientific

Sourced in United States, China, United Kingdom, Germany, Australia, Japan, Canada, Italy, France, Switzerland, New Zealand, Brazil, Belgium, India, Spain, Israel, Austria, Poland, Ireland, Sweden, Macao, Netherlands, Denmark, Cameroon, Singapore, Portugal, Argentina, Holy See (Vatican City State), Morocco, Uruguay, Mexico, Thailand, Sao Tome and Principe, Hungary, Panama, Hong Kong, Norway, United Arab Emirates, Czechia, Russian Federation, Chile, Moldova, Republic of, Gabon, Palestine, State of, Saudi Arabia, Senegal

Fetal Bovine Serum (FBS) is a cell culture supplement derived from the blood of bovine fetuses. FBS provides a source of proteins, growth factors, and other components that support the growth and maintenance of various cell types in in vitro cell culture applications.

Facsaria 2 by BD

Sourced in United States, Germany, United Kingdom, Japan, Belgium, China, Canada, Italy, France, South Sudan, Singapore, Australia, Denmark, Uruguay

The FACSAria II is a high-performance cell sorter produced by BD. It is designed for precision cell sorting and analysis. The system utilizes flow cytometry technology to rapidly identify and separate different cell populations within a sample.

Nanodrop by Thermo Fisher Scientific

Sourced in United States, Germany, United Kingdom, China, France, Japan, Canada, Italy, Belgium, Australia, Denmark, Spain, Sweden, India, Finland, Switzerland, Poland, Austria, Brazil, Singapore, Portugal, Macao, Netherlands, Taiwan, Province of China, Ireland, Lithuania

The NanoDrop is a spectrophotometer designed for the quantification and analysis of small volume samples. It measures the absorbance of a sample and provides accurate results for DNA, RNA, and protein concentration measurements.

What are the unique properties of cyclic peptides?

Cyclic peptides have a circular structure formed by a covalent bond between the amino and carboxyl termini. This confers several advantageous properties, including increased stability, resistance to proteolytic degradation, and improved cell permeability compared to linear peptides. The cyclic structure also gives cyclic peptides unique physicochemical and pharmacological activities, making them valuable for various therapeutic applications.

How are cyclic peptides found and produced?

What are some common applications of cyclic peptides?

How can PubCompare.ai assist in cyclic peptide research?

PubCompare.ai's AI-driven platform can accelerate cyclic peptide research in several ways. First, it allows researchers to screen protocol literature more efficiently, helping them quickly locate relevant protocols from published literature, pre-prints, and patents. Second, the platform's AI-powered analysis can pinpointe critical insights, enabling researchers to identify the most effective protocols related to cyclic peptides for their specific research goals. This helps ensure reproducibility and accuracy by highlighting key differences in protocol effectiveness, allowing researchers to choose the best option.

What are some common challenges in working with cyclic peptides?

One of the main challenges in working with cyclic peptides is their relatively poor solubility and cell permeability compared to linear peptides. This can make it difficult to formulate and deliver cyclic peptides effectively. Additionally, the synthesis of cyclic peptides can be more complex and time-consuming than linear peptides. Researchers may also face challenges in consistently producing cyclic peptides with the desired purity and tytpical properties.

More about "Cyclic Peptides"

Cyclic peptides are a class of peptides with a unique circular structure formed by a covalent bond between the amino and carboxyl termini.
These macrocyclic molecules exhibit distinctive physicochemical properties and pharmacological activities, making them invaluable in various therapeutic applications.
Naturally occurring cyclic peptides can be found in microorganisms, plants, and animals, and they can also be synthetically produced.
The cyclic structure of these peptides confers increased stability, resistance to proteolytic degradation, and improved cell permeability compared to their linear counterparts.
This enhanced stability and cell-penetrating ability make cyclic peptides valuable in the development of novel drugs, diagnostics, and biotechnological tools.
Researchers studying cyclic peptides can leverage the power of AI-driven platforms like PubCompare.ai to accelerate their research.
These platforms can help scientists easily locate relevant protocols from literature, preprints, and patents, and utilize AI-powered comparisons to identify the best protocols and products.
Specialized equipment like the J-815 CD spectrometer, Milli-Q water purification system, J-815 spectropolarimeter, J-810 spectropolarimeter, Voyager workstation, 415 autosampler, and FACSAria II flow cytometer can be employed in the analysis and characterization of cyclic peptides.
Techniques such as circular dichroism (CD) spectroscopy, using the J-815 spectrometer, can provide insights into the secondary structure and conformational properties of these macrocyclic molecules.
Whether you're a researcher interested in developing novel cyclic peptide-based therapeutics, diagnostics, or biotechnological applications, or a scientist exploring the fundamental properties of these unique peptides, the insights and tools available can help you unlock new frontiers in cyclic peptide research and innovation.