Increase in ADP‐ribosyltransferase activity of rat T lymphocyte alloantigen RT6.1 by a single amino acid mutation

FEBS Letters

Hoshino, Shin-ichi¹  Katada, Toshiaki¹  Maehama, Tomohiko¹ 

[1] Department of Physiological Chemistry, Faculty of Pharmaceutical Sciences, University of Tokyo, Tokyo 113, Japan

关键词: ADP-ribosylation;    ADP-ribosyltransferase;    Rat T lymphocyte alloantigen;    GPI;    glycosylphosphatidylinositol;    MBP;    maltose-binding protein;   

DOI  :  10.1016/0014-5793(96)00568-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

A family of glycosylphosphatidylinositol-linked ADP-ribosyltransferases, of which cDNAs were cloned from various mammalian cells, possess a common Glu-rich motif (EEEVLIP) near their carboxyl termini. Although the first Glu in the common motif is replaced by Gln (Q²⁰⁷EEVLIP) in rat T lymphocyte alloantigens RT6.1 and RT6.2, the two RT6s appear to have both activities of NAD⁺ glycohydrolase and ADP-ribosyltransferase to a lesser extent. To investigate the significance of the Glu-rich motif in the two enzyme activities, we produced a mutant RT6.1 (Q207E), in which Gln²⁰⁷ was replaced by Glu, together with wild-type RT6s, in Escherichia coli. Kinetic analysis revealed that there were no marked differences in the V _max and K _m values of NAD⁺ glycohydrolases among the three recombinant proteins. The recombinant RT6.1 and RT6.2 displayed extremely low auto-ADP-ribosylation, although the latter modification was somewhat higher than the former. In contrast, much greater auto-modification was observed for the Q207E mutant. Moreover, the mutant could effectively ADP-ribosylate agmatine as a substrate. Thus, the single amino acid mutation of RT6.1 caused a marked increase in its ADP-ribosyltransferase activity, indicating that the Glu-rich motif near the carboxy terminus plays an important role in the enzyme activity.

【 授权许可】

Unknown