ON THE REACTIVITY OF CARBOXYL GROUPS OF RIBONUCLEASE‐A IN ANHYDROUS METHANOL

A. Seetharama Acharya, Paul J. Vithayathil

Research output: Contribution to journalArticle

8 Scopus citations

Abstract

The esterification of Ribonuclease‐A in methanol/0.1 M hydrochloric acid has been studied by measuring the decrease in the number of titratable groups of the protein and estimating the amount of methanol incorporated. Esterification of nearly five of the 11 free carboxyl groups of the protein resulted in almost complete inactivation of the enzyme. The initial products of esterification have been chromatographed on Amberlite columns, and five partially active methyl ester derivatives of Ribonuclease‐A have been isolated. The dimethyl ester, the initial product of esterification with reduced catalytic activity, has the carboxyl groups of Glu‐49 and Asp‐53 modified. Even in the non‐aqueous solvent, as in the native structure of the protein in aqueous solution, these carboxyl groups are the fast reacting ones. Subsequently, the esterification reaction appears to proceed preferentially at the C‐terminal region of the molecule. Comparison of the reactivities of carboxyl groups of Ribonuclease‐A in acidic methanol to that known in aqueous solutions (with carbodiimides) suggests that the structure of Ribonuclease‐A in the non‐aqueous solvent resembles, at least in part, the structure in aqueous environment.

Original languageEnglish (US)
Pages (from-to)207-219
Number of pages13
JournalInternational Journal of Peptide and Protein Research
Volume7
Issue number3
DOIs
StatePublished - May 1975

ASJC Scopus subject areas

  • Biochemistry

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