32p ppi

Manufactured by PerkinElmer

[32P]-PPi is a radioactive isotope of phosphorus that is used as a tracer in various biochemical and molecular biology applications. It is a stable compound that is typically used to label and detect specific biomolecules, such as nucleic acids, proteins, and other phosphate-containing compounds.

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Lab products found in correlation

Etidronate, by Merck Group (2 mentions) Pamidronate, by Merck Group (2 mentions) Imidodiphosphate, by Merck Group (2 mentions) Ddctp, by GE Healthcare (2 mentions) Chlodronate, by Merck Group (2 mentions) 3 azido 2 3 dideoxythymidine triphosphate azttp, by TriLink (2 mentions) Arabinofuranosylcytosine triphosphate aractp, by TriLink (2 mentions) Gemcitabine dfdctp, by Jena Biosciences (2 mentions) α 35s datp, by PerkinElmer (2 mentions) Polyethyleneimine pei cellulose thin layer chromatography plates, by Merck Group (2 mentions) α 32p dctp, by PerkinElmer (2 mentions) Proline, by Merck Group (1 mentions) Adenosine triphosphate (atp), by Merck Group (1 mentions) γ 32p atp, by PerkinElmer (1 mentions)

4 protocols using 32p ppi

Enzymatic Assay for Pyrophosphate

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Proline, ATP, and other reagents including metal salts and buffers were from Sigma (>99% pure)). [γ−³²P]-ATP and [³²P]-PP_i were from Perkin Elmer. Primers for site-directed mutagenesis and PCR were from Integrated DNA Technologies. Inorganic pyrophosphatase (PP_iase) and reagents for in vitro transcription were from Fisher Scientific.

Zajac J., Anderson H., Adams L., Wangmo D., Suhail S., Almen A., Berns L., Coerber B., Dawson L., Hunger A., Jehn J., Johnson J., Plack N., Strasser S., Williams M., Bhattacharyya S, & Hati S. (2020). Effects of Distal Mutations on Prolyl-adenylate Formation of Escherichia coli Prolyl-tRNA Synthetase. The protein journal, 39(5), 542-553.

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Human pol β purification and assay

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Human pol β was expressed and purified^{44 (link)}. Chlodronate, etidronate, imidodiphosphate, pamidronate, and pyrophosphate were from Sigma-Aldrich. The β,γ-imido modified nucleoside triphosphate analog, 2′-deoxyguanosine-5′-[(β,γ)-imido]triphosphate (dGMPPNP), was from Jena Bioscience. Chain terminating nucleoside triphosphates; ddCTP was from GE Healthcare, 3′-azido-2′,3′-dideoxythymidine triphosphate (AZTTP) and arabinofuranosylcytosine triphosphate (araCTP) were from Trilink Biotechnologies, and gemcitabine (dFdCTP) was obtained from Jena Bioscience. [α-³⁵S]dATP, [α-³²P]dCTP, and [³²P]PP_i were from Perkin Elmer. Polyethyleneimine (PEI) cellulose thin layer chromatography (TLC) plates containing a fluorescent indicator were purchased from EMD Millipore.

Shock D.D., Freudenthal B.D., Beard W.A, & Wilson S.H. (2017). Modulating the DNA polymerase β reaction equilibrium to dissect the reverse reaction. Nature chemical biology, 13(10), 1074-1080.

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Human pol β purification and assay

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Shock D.D., Freudenthal B.D., Beard W.A, & Wilson S.H. (2017). Modulating the DNA polymerase β reaction equilibrium to dissect the reverse reaction. Nature chemical biology, 13(10), 1074-1080.

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Monitoring Amino Acid Activation by ATP/PP_i Exchange

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Amino acid activation was monitored using ATP/PP_i exchange as previously described [60 ]. Reactions were performed at 37°C in 100 mM HEPES pH 7.2, 30 mM KCl, 10 mM MgCl₂, 2 mM NaF, 2 mM ATP, 2 mM [³²P]-PP_i (Perkin Elmer), 90 μM alanine, 160 nM AlaRS, and DMSO or aaRS inhibitor. At increasing time points, aliquots of the reaction mixture were quenched in a charcoal solution containing 1% activated charcoal, 5.6% HClO₄, and 75 mM PP_i. Quenched reactions were vacuum filtered on to 3MM Whatman filter discs, washed three times with 5 mL of water and once with 5 mL of ethanol. After drying the filter discs, charcoal-bound radiolabeled ATP was quantified on a scintillation counter. Relative endpoint amino acid activation was determined by comparing the inhibitor-treated enzymes to their respective DMSO control samples.

Kelly P., Hadi-Nezhad F., Liu D.Y., Lawrence T.J., Linington R.G., Ibba M, & Ardell D.H. (2020). Targeting tRNA-synthetase interactions towards novel therapeutic discovery against eukaryotic pathogens. PLoS Neglected Tropical Diseases, 14(2), e0007983.

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