Subunit separation in reversed micelle system reveals the existence of active centers both on light and heavy γ‐glutamyltransferase subunits

FEBS Letters

Chernov, Nikolai N.¹  Klyachko, Natalya L.²  Kabanov, Alexander V.²  Levashov, Andrey V.²  Nametkin, Sergey N.² 

[1] Department of Biochemistry, Patrice Lumumba University, Moscow, Russia;Laboratory of Micellar Enzymology, Division of Chemical Enzymology, Department of Chemistry, Moscow State University, Moscow, Russia

关键词: γ-Glutamyltransferase;    Protein subunit;    Micellar enzymology;    Reversed micelle;    Supra-macromolecular structure;    γ-GT;    γ-glutamyltransferase;    AOT;    Aerosol OT (sodium bis(2-ethylhexyl)sulfosuccinate);    NA;    (4-nitro)aniline;    CNA;    (3-carboxy-4-nitro)aniline;    GluNA;    γ-glutamyl(4-nitro)anilide;    GluCNA;    γ-glutamyl(3-carboxy-4-nitro)anilide;    AT-125;    L-(αS;    5S)-α-amino-3-chloro-4;    5-dihydro-5-isoxazoleacetic acid;    GlyGly;    glycylglycine;    RM;    reversed micelle;    PAGE;    polyacrylamide gel electrophoresis;   

DOI  :  10.1016/0014-5793(91)81388-O

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Regulation of supra-macromolecular composition and catalytic activity of a heterodimeric enzyme, γ-glutamyltransferase, in the system of Aerosol OT (sodium bis(2-ethylhexyl) sulfosuccinate) reversed micelles in octane were studied. Variation of the surfactant hydration degree (parameter, determining dimensions of the polar inner cavity of the micelle) causes a reversible dissociation of the enzyme to light and heavy subunits. Both enzyme subunits possess catalytic activity. The light and heavy subunits of the enzyme were separated on a preparative scale in a reversed micelle system using ultracentrifugation. The active centers of γ-glutamyltransferase were studied using its irreversible inhibitor — AT-125 (L·(αS.5S)-α-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid). Separation of the γ-glutamyltransferase subunits results in the ‘opening’ of a new active center located at the heavy subunit. In the dimer form of the enzyme this center is masked and it is not accessible to both substrate and inhibitor molecules.

【 授权许可】

Unknown