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Isopropyl β d thiogalactopyranoside

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Isopropyl β-D-thiogalactopyranoside is a synthetic chemical compound used as an inducer in recombinant protein expression systems. It is a structural analog of the natural lactose metabolite allolactose, which serves as the natural inducer of the lac operon in Escherichia coli.

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Lab products found in correlation

Escherichia coli strain bl21 gold de3, by New England Biolabs (2 mentions) Dpni, by New England Biolabs (2 mentions) Protamine sulfate, by MP Biomedicals (2 mentions) Lb broth, by Research Products International (2 mentions) Kanamycin, by Research Products International (2 mentions) Wizard plus sv miniprep dna purification system, by MP Biomedicals (2 mentions) Vivaspincentrifugal concentrator, by MP Biomedicals (2 mentions) Deuterium oxide, by Cambridge Isotopes (2 mentions) Vivaspin 20 spin filters, by Cytiva (2 mentions) Lysozyme, by Merck Group (2 mentions) Dithiothreitol (dtt), by Merck Group (2 mentions) Agarose, by Research Products International (1 mentions) Pfu turbo dna polymerase, by Agilent Technologies (1 mentions) Trimethylchlorosilane tmcs, by Merck Group (1 mentions) Pyrophosphatase, by Merck Group (1 mentions) Neuraminic acid, by Merck Group (1 mentions) Imidazole, by Merck Group (1 mentions) Lactate dehydrogenase, by Merck Group (1 mentions) Phosphoenolpyruvate, by Merck Group (1 mentions) Udp glcnac, by Merck Group (1 mentions)

Spelling variants of this product

Isopropyl β-D-1-thiogalactopyranoside (6 citations) Isopropyl-β-D-thiogalactopyranoside (IPTG) (5 citations)

9 protocols using isopropyl β d thiogalactopyranoside

1

Enzymatic Synthesis of GDP-Glycero-Manno-Heptose

Cited 1 time
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Unless otherwise noted, all chemicals used in this study were obtained from Sigma-Aldrich, Carbosynth, or GE Healthcare Biosciences. The bacterial growth medium, agarose and isopropyl-β-d-thiogalactopyranoside (IPTG) were purchased from Research Products International (RPI). Escherichia coli strain BL21-Gold (DE3) and the restriction enzyme DpnI were obtained from New England Biolabs. PfuTurbo DNA polymerase was purchased from Agilent. DNA sequencing was conducted by Eton Biosciences Inc. GDP-d-glycero-α-d-manno-heptose (1) was enzymatically synthesized and purified as described previously (9 (link)).
Xiang D.F., Ghosh M.K., Riegert A.S., Thoden J.B., Holden H.M, & Raushel F.M. (2022). Bifunctional Epimerase/Reductase Enzymes Facilitate the Modulation of 6-Deoxy-Heptoses Found in the Capsular Polysaccharides of Campylobacter jejuni. Biochemistry, 62(1), 134-144.
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2

5-Nitrovanillate Synthesis and Characterization

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5-nitrovanillate (5-NV)
was synthesized according
to published procedures.24 (link)Pfu
Turbo DNA polymerase and the Escherichia coli strains BL21(DE3) and XL1-blue cells were obtained from Stratagene.
The restriction enzyme DpnI was purchased from New England BioLabs
and oligonucleotides were obtained from Integrated Data Technology
through the Gene Technology Laboratory at Texas A&M University.
Kanamycin, isopropyl β-D-thiogalactopyranoside (IPTG), and LB
broth were acquired from Research Products International Corp. Protamine
sulfate, Wizard Plus SV Miniprep DNA purification system, and Vivaspin
centrifugal concentrator (10 000 MWCO) were obtained from MP
Biomedicals LLC., Promega, and Fisher Scientific, respectively.
Vladimirova A., Patskovsky Y., Fedorov A.A., Bonanno J.B., Fedorov E.V., Toro R., Hillerich B., Seidel R.D., Richards N.G., Almo S.C, & Raushel F.M. (2015). Substrate Distortion and the Catalytic Reaction Mechanism of 5-Carboxyvanillate Decarboxylase. Journal of the American Chemical Society, 138(3), 826-836.
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3

Synthesis and Characterization of Isotopically Labeled Compounds

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Chemicals, unless otherwise indicated, were purchased from Sigma-Aldrich. Isopropyl β-D-thiogalactopyranoside and all buffers used in this study were purchased from Research Products International (Mt. Prospect, IL). Imidazole and Luria broth were purchased from bio-WORLD (Dublin, OH). Isotopically labeled H218O2 (90% 18O) was purchased from ICON Isotopes (Summit, NJ). Perdeuterated fatty acids were purchased from CDN Isotopes (Pointe-Claire, QC). Protiated fatty acids, N,O-bis(trimethylsilyl)-trifluoroacetamide (BSTFA), and trimethylchlorosilane (TMCS) (99:1) were purchased from Supelco (Bellefonte, PA). Radical clock substrates were synthesized according to published procedures.33 ,34
Hsieh C.H., Huang X., Amaya J.A., Rutland C.D., Keys C.L., Groves J.T., Austin R.N, & Makris T.M. (2017). The Enigmatic P450 Decarboxylase OleT Is Capable of, but Evolved To Frustrate, Oxygen Rebound Chemistry. Biochemistry, 56(26), 3347-3357.
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4

In Vitro Protein Binding Assay for SOD1 and Parkin

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His-tagged SOD1 WT or mutants, glutathione S-transferase (GST), and GST-tagged parkin were individually expressed in E. coli BL21 or Arctic Express cells and induced with isopropyl β-D-thiogalactopyranoside (IPTG, 0.1 mM for GST-tagged proteins or 1 mM for His-tagged proteins; Research Products International) as described [44 (link)]. GST-tagged proteins were purified using glutathione-agarose beads as we described [45 (link), 46 (link)], and His-tagged proteins were purified using Ni-NTA-agarose beads and dialyzed in a buffer containing 100 μM each of CuSO4 and ZnSO4 as described [47 (link)]. In vitro binding assays were performed as described [48 (link)] by incubation of GST-tagged parkin or GST immobilized on glutathione-agarose with purified His-tagged SOD1 WT or mutants, and bound proteins were detected by immunoblotting analysis.
Yung C., Sha D., Li L, & Chin L.S. (2015). Parkin Protects Against Misfolded SOD1 Toxicity by Promoting Its Aggresome Formation and Autophagic Clearance. Molecular neurobiology, 53(9), 6270-6287.
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5

Enzymatic Production of Sialic Acid Derivatives

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Lysogeny broth (LB), isopropyl β-d-thiogalactopyranoside (IPTG), NAD+, and NADH were
purchased from Research Products International. The protease inhibitor
cocktail, lysozyme, DNase I, UDP-GlcNAc, ManNAc, neuraminic acid,
uridine 5′-diphosphate (UDP), cytidine 5′-triphosphate
(CTP), cytidine 3′,5′-cyclic monophosphate, phosphoenolpyruvate
(PEP), pyruvate kinase, lactate dehydrogenase, sialic acid aldolase,
pyrophosphatase, kanamycin, dithiothreitol (DTT), imidazole, and HEPES
were obtained from Sigma-Aldrich. Ammonium bicarbonate, 2-mercaptoethanol,
KCl, MnCl2, and MgCl2 were acquired from Sigma-Aldrich.
Vivaspin 20 spin filters and HisTrap and HiTrap Q HP columns were
obtained from Cytiva. The 10 kDa Nanosep spin filters were purchased
from Pall Corp. (Port Washington, NY). Deuterium oxide was acquired
from Cambridge Isotope Laboratories Inc., and 18O-labeled
water (98%) was obtained from Medical Isotopes Inc.
Ghosh M.K, & Raushel F.M. (2024). Biosynthesis of UDP-α-N-Acetyl-d-mannosaminuronic Acid and CMP-β-N-Acetyl-d-neuraminic Acid for the Capsular Polysaccharides of Campylobacter jejuni. Biochemistry, 63(5), 688-698.
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6

Lipid Preparation for Biophysical Studies

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The list of chemicals and their suppliers are: brain sulfatides, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE) (Avanti-Lipids), N-dodecylphosphocholine (DPC) (Anatrace), cholesterol (Sigma), and isopropyl β-D-thiogalactopyranoside (Research Products International). All other chemicals were of analytical grade.
Song W., Gottschalk C.J., Tang T.X., Biscardi A., Ellena J.F., Finkielstein C.V., Brown A.M, & Capelluto D.G. (2020). Structural, in silico, and functional analysis of a Disabled-2-derived peptide for recognition of sulfatides. Scientific Reports, 10, 13520.
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7

Synthesis and Purification of GDP-D-glycero-α-D-manno-heptose

Cited 1 time
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All
materials used in this study were obtained
from Sigma-Aldrich, Carbosynth, or GE Healthcare Biosciences, unless
otherwise stated. Lysogeny broth (LB) and isopropyl-β-d-thiogalactopyranoside (IPTG) were purchased from Research Products
International. HisTrap columns and Vivaspin 20 spin filters were obtained
from Cytiva. The 3 and 10 kDa Nanosep spin filters were purchased
from Pall Corporation (Port Washington, NY). Deuterium oxide was purchased
from Cambridge Isotope Laboratories, Inc. Escherichia coli strain BL21-Gold (DE3) was obtained from New England Biolabs. α-Ketoglutarate
was purchased from AK Scientific (Union City, CA). Ultraviolet spectra
were recorded on a SpectraMax340 (Molecular Devices) ultraviolet–visible
plate reader using 96-well Greiner plates. 1H NMR and 1H–1H COSY spectra were recorded on a Bruker
Avance III 400 MHz system equipped with a broadband probe and sample
changer. Mass spectrometry data were collected on a Thermo Scientific
Q Exactive Focus system run in the negative-ion mode. GDP-d-glycero-α-d-manno-heptose (5) was synthesized and purified as described
previously.20 (link)
Xiang D.F., Xu M., Ghosh M.K, & Raushel F.M. (2023). Metabolic Pathways for the Biosynthesis of Heptoses Used in the Construction of Capsular Polysaccharides in the Human Pathogen Campylobacter jejuni. Biochemistry, 62(21), 3145-3158.
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8

Purification of Membrane Proteins

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Yeast extract, tryptone, and glycerol were purchased from Fisher Scientific. Sodium cholate of >99% purity was obtained from Anatrace. Isopropyl-β-D-thiogalactopyranoside (IPTG), carbenicillin (disodium salt) and δ-aminolevulinic acid were from Research Products International Corp. DE52 media (preswollen diethylaminoethyl cellulose) was from Whatman. Bio-Beads SM-2 and Hydroxyapatite resins were from Bio-Rad. Dithiothreitol, Tergitol NP-10, nonaethylene glycol monododecyl ether (Brij), Cymal-5, Reactive Red 120 type 300-CL, Octyl Sepharose CL-4B, 2,6-ditertbutyl-4-methylphenol butylated hydroxytoluene (BHT), Trizma base, and lysozyme were from Sigma. DNase and the protease inhibitor cocktail were purchased from Roche Diagnostics Corp. (Indianapolis). 50% poly-ethylene glycol (PEG) 3350 solution, 10% w/v of Anapoe-C10E9 solution, 1 M magnesium acetate solution, 1 M sodium cacodylate solution, and 1 M HCl solution were purchased from Hampton Research (California).
Yang Y., Bu W., Im S., Meagher J., Stuckey J, & Waskell L. (2018). Structure of cytochrome P450 2B4 with an acetate ligand and an active site hydrogen bond network similar to oxyferrous P450cam. Journal of inorganic biochemistry, 185, 17-25.
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9

Cloning and Protein Expression Protocol

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All chemicals, buffers, and purification reagents were purchased from Sigma-Aldrich unless otherwise specified. Pfu Turbo DNA polymerase and the E. coli strains BL21 (DE3) and XL1-blue cells were obtained from Stratagene. The restriction enzyme DpnI was purchased from New England BioLabs and oligonucleotides were obtained from Integrated Data Technology through the Gene Technology Laboratory at Texas A&M University. Kanamycin, isopropyl β-D-thiogalactopyranoside (IPTG), and LB broth were acquired from Research Products International Corp. Protamine sulfate, Wizard Plus SV Miniprep DNA purification system, and Vivaspin centrifugal concentrator (10 000 MWCO) were obtained from MP Biomedicals LLC., Promega, and Fisher Scientific, respectively. Chromatographic gel filtration columns were purchased from GE Healthcare. 2-Nitroresorcinol was purchased from Sigma Aldrich.
Sheng X., Patskovsky Y., Vladimirova A., Bonanno J.B., Almo S.C., Himo F, & Raushel F.M. (2018). Mechanism and Structure of γ-Resorcylate Decarboxylase. Biochemistry, 57(22), 3167-3175.
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