Modification of the acceptor binding site of γ-glutamyl transpeptidase by the diazonium derivatives of p-aminohippurate and p-aminobenzoate Academic Article uri icon

Overview

MeSH Major

  • Phospholipid Transfer Proteins

abstract

  • Hippurate and maleate have been shown to bind to the aminoacylglycine (acceptor) binding site of γ-glutamyl transpeptidase, thereby stimulating the hydrolysis of γ-glutamyl compounds at the expense of transpeptidation (Thompson, G. A., and Meister, A. (1979) J. Biol. Chem. 254, 2956-2960; Thompson, G. A., and Meister, A. (1980) J. Biol. Chem. 255, 2109-2113). It has now been found that a number of benzoate derivatives also bind and modulate rat kidney transpeptidase, as indicated by their ability to enhance the rate of inactivation of transpeptidase by the glutamine antagonist l-(αS, 5S)-α-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid (AT-125). Furthermore, rapid loss of transpeptidase activity results upon preincubation of the enzyme with the diazonium derivatives of p-aminohippurate and p-aminobenzoate. The modified enzyme can still hydrolyze γ-glutamyl substrates but is no longer modulated by hippurate and maleate. Loss of transpeptidase activity was not associated with incorporation of radioactive label from diazotized [14C]p-aminohippurate. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the modified enzyme revealed a nondissociable species, Mr 68,000, shown to result from crosslinking of the two subunits of transpeptidase (Mr 46,000 and 22,000, respectively). The crosslinking of the subunits paralleled the extent of inactivation of transpeptidation activity and both crosslinking and inactivation were prevented by treatment with the diazotized derivatives in the presence of either hippurate or maleate. These and other data indicate that the diazonium derivatives of p-aminohippurate and p-aminobenzoate interact with the acceptor binding site and produce a stable bond between amino acid residues in the vicinity of this site which, thus, appears to be located in the intersubunit contact region. © 1982.

publication date

  • January 1982

Research

keywords

  • Academic Article

Identity

Digital Object Identifier (DOI)

  • 10.1016/0003-9861(82)90262-4

Additional Document Info

start page

  • 719

end page

  • 26

volume

  • 216

number

  • 2