The second nucleophile molecule binds to the acyl‐enzyme–nucleophile complex in α‐chymotrypsin catalysis: Kinetic evidence for the interaction

Mikhail Y. GOLOLOBOV, Valentin M. STEPANOV, Tatjana L. VOYUSHINA, Patrick ADLERCREUTZ

    Research output: Contribution to journalArticlepeer-review

    Abstract

    α‐Chymotrypsin‐catalyzed acyl tranfer was studied using three acyl‐group donors (Mal‐l‐Ala‐l‐Ala‐l‐PheOMe, Bz‐l‐TyrOEt and Ac‐l‐TrpOEt; Mal, maleyl; Bz, benzoyl; OMe, methyl ester; OEt, ethyl ester) and a series of amino‐acid amides. Most of the reactions studied can be described by the simplest kinetic model without the nucleophile binding to the acyl‐enzyme. The α‐chymotrypsin‐catalyzed transfer of the Mal‐l‐Ala‐l‐Ala‐l‐Phe group to the amides of L‐Phe and L‐Tyr showed a linear dependence of the partition constant, p, on the nucleophile concentration which can be interpreted by the hydrolysis of the acyl‐enzyme–nucleophile complex. The α‐chymotrypsin‐catalyzed transfer of the Bz‐l‐Tyr and Ac‐l‐Trp groups to several amino‐acid amides showed unusual behavior which can be interpreted by the kinetic model involving formation of a complex of the acyl‐enzyme with two nucleophile molecules. These observations can explain the conflicting conclusions concerning the kinetics of α‐chymotrypsin‐catalyzed acyl transfer evident in previous studies.

    Original languageEnglish
    Pages (from-to)955-963
    Number of pages9
    JournalEuropean Journal of Biochemistry
    Volume217
    Issue number3
    DOIs
    Publication statusPublished - 1993 Jan 1

    Subject classification (UKÄ)

    • Biocatalysis and Enzyme Technology

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