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. 1986 Apr 23;357(1):191-8.
doi: 10.1016/s0021-9673(01)95820-0.

Highly sensitive assay for gamma-glutamyltranspeptidase activity by high-performance liquid chromatography with electrochemical detection

K Kiuchi, K Kiuchi, T Nagatsu, A Togari, H Kumagai

Highly sensitive assay for gamma-glutamyltranspeptidase activity by high-performance liquid chromatography with electrochemical detection

K Kiuchi et al. J Chromatogr. .

Abstract

A highly sensitive assay for gamma-glutamyltranspeptidase activity involving high-performance liquid chromatography (HPLC) with electrochemical detection was devised. gamma-Glutamyl-DOPA, a new synthetic dipeptide, which consists of naturally occurring amino acids, was found to be a good substrate for gamma-glutamyltranspeptidase purified from Proteus mirabilis. Enzymatically formed DOPA was adsorbed on an aluminium oxide column, eluted with 0.5 M hydrochloric acid and determined by HPLC with electrochemical detection. The sensitivity limit of this method was 0.5 pmol of DOPA formed. Some properties of gamma-glutamyltranspeptidase purified from P. mirabilis were investigated using gamma-glutamyl-DOPA as a substrate. In the presence of 0.15 M glycylglycine, the KM value of the enzyme for gamma-glutamyl-DOPA was 0.013 mM, and the maximum velocity was 247 nmol/min per mg protein. This method was applied to the assay of the enzymatic activity in human serum.

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