Scientific Reports of the Faculty of Agriculture, Okayama University
Published by the Faculty of Agriculture, Okayama University
ONLINE ISSN : 2186-7755

好熱好酸性アーキア Sulfolobus tokodaii 由来シスタチオニン γ-シンターゼの精製及び性質検討

Shinozaki, Mai
Yanagitani, Masahiko
Kaneda, Shouichirou
Kudou, Daizou
Endou, Yuuichi
Kuramitsu, Seiki
Abstract
The gene encoding a cystathionine γ-synthase from Sulfolobus tokodaii was cloned and expressed in Escherihia coli Rosetta-gami (DE3). Cystathionine γ-synthase [EC 2. 5. 1. 48] from Sulfolobus tokodaii (stCGS) was purified by heat treatment, DEAE- Toyopearl 650M and Sephacryl S-300 column chromatographies from E. coli transformants. stCGS shows optimum activity at pH 7.0, and is stable between pH5.0 and pH9.0. The optimum temperature of stCGS is above 100℃, and the enzyme showed the remaining activity of almost 100% up to 70℃. The K(m) and V(max) with O-phospho-L- homoserine as a substrate are 0.82 mM and 2.42 U/mg. To analyze the role of Phe 97 in the active site of stCGS, we constructed F97Y, R99C, and F97Y-R99C mutant enzymes. Although native stCGS has no activity toward l-methionine, F97Y mutant enzyme gained the elimination activity toward L-methionine.
Keywords
cystathionine γ-synthase
pyridoxal 5’-phosphate
thermoacidophilic archaea
Sulfolobus tokodaii
ISSN
0474-0254 
NCID
AN00033029