Identification of cysteine as the reactive group in pyruvate kinase alkylated by 5-chloro-4-oxopentanoic acid.

doi:10.1042/BJ1590213

Paper

Identification of cysteine as the reactive group in pyruvate kinase alkylated by 5-chloro-4-oxopentanoic acid.

Published Nov 1, 1976 · R. Chalkley, D. Bloxham

The Biochemical journal

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5

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0

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Abstract

4-Hydroxypentanoic acid alanine thioether was synthesized and characterized by n.m.r. spectroscopy. This derivative corresponded to the modified amino acid obtained by allowing 5-chloro-4-oxo[3,5-3H]pentanoic acid to react with rabbit muscle pyruvate kinase. Performic acid oxidation of 4-oxo[3,5-3H]pentanoic acid alanine thioether in pyruvate kinase gave [3H]succinate (67%) and [3H]carboxymethylcysteine (33%) as expected. Evidence is presented to show that NaBH4 reduction followed by periodate oxidation and analysis of radioactive formaldehyde production may provide a convenient method for distinguishing between thiol and amino alkylation by halogenomethyl ketone compounds. Peptide 'mapping' confirms that the modification by 5-chloro-4-oxopentanoic acid occurs primarily at one region of pyruvate kinase.

In Vitro Study

Study Snapshot

5-chloro-4-oxopentanoic acid modifies pyruvate kinase primarily at one region, with cysteine as the reactive group, and NaBH4 reduction and periodate oxidation can distinguish between thiol and amino alkylation.

PopulationOlder adults (50-71 years)

Sample size24

MethodsObservational

OutcomesBody Mass Index projections

ResultsSocial networks mitigate obesity in older groups.

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Paper

Identification of cysteine as the reactive group in pyruvate kinase alkylated by 5-chloro-4-oxopentanoic acid.

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References

Citations

Modification of two essential cysteines in rabbit muscle pyruvate kinase by the guanine nucleotide analogue 5'[p-(fluorosulfonyl) benzoyl] guanosine.

The inactivation of rabbit muscle pyruvate kinase by 5'-FSBG involves two essential cysteines, with the restoration of activity correlated to the regeneration of two free sulfhydryl groups after treatment with dithiothreitol.

Isolation and sequence determination of a peptide located in or near the active site of bovine muscle pyruvate kinase.

A 34-amino acid peptide with -trinitrophenylated lysine was identified in the active site of bovine muscle pyruvate kinase.

Use of methanethiolation to investigate the catalytic role of sulphydryl groups in rabbit skeletal muscle pyruvate kinase.

Methanethiosulphonate reveals that only a single thiol group participates in the catalytic event of rabbit skeletal muscle pyruvate kinase, with its role in maintaining enzyme structure also being studied.

[47] Chloromethyl ketone derivatives of fatty acids

Chloromethyl ketone fatty acid analogs are useful for designing specific enzyme inhibitors, with potential applications in enzymology involving fatty acid binding and metabolism.

5-Aminolevulinic Acid Dehydratase

Groups I and II of 5-aminolevulinic acid dehydratase are highly reactive with Nbs2, forming an intramolecular disulphide bond, while group III maintains conformation in the native enzyme.