ISHS Acta Horticulturae 688: IV International Symposium on Edible Alliaceae
PURIFICATION, CHARACTERIZATION AND CLONING OF ONION GAMMA-GLUTAMYL TRANSPEPTIDASE |
| Authors: | M. Shaw, M. Pither-Joyce, M. McManus, J. McCallum |
| Keywords: | gamma-glutamyl transpeptidase, flavour, onion |
| Abstract:
Gamma-glutamyl transpeptidase (E.C. 2.3.2.2; GGT) catalyses the hydrolysis of gamma-glutamyl linkages of gamma-glutamyl peptides and the transfer of the gamma-glutamyl group to a large number of amino acids and peptides. In onion, GGT is believed to catalyse the last step in the biosynthesis of flavour precursors by cleaving the gamma-glutamyl moiety off gamma-glutamyl alk(en)yl cysteine sulfoxides. Plant GGTs are poorly characterized with the purification of enzymes from different plant species resulting in contradicting data regarding molecular masses, subunit composition, cellular localization and biochemical characteristics. We obtained cDNA clones from root and leaf with homology to plant GGTs by RT-PCR and RACE. Sequence analysis showed an N-terminal signal peptide and processing site similar to Arabidopsis GGT. We purified an enzyme from sprouting bulbs that exhibit GGT activity with the artificial substrate gamma-Glu-p-nitroaniline. The bulb enzyme is a membrane associated glycoprotein and probably has heterodimer structure with the larger subunit of 38.9 kDa molecular mass. We confirmed homology of this purified subunit with the cloned cDNA by N-terminal sequencing. Kinetic studies suggest that this enzyme is unlikely to catalyse hydrolysis of gamma-glutamyl ACSOs but could catalyse transpeptidation of precursors such as S-methyl glutathione. | |
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