Purification and Characterization of Thermostable Glycerol Kinase from Thermus flavus

元データ 1997-04-25 社団法人日本生物工学会

概要

Glycerol kinase (EC 2.7.1.30; ATP : glycerol 3-phosphotransferase) was purified from Thermus flavus, by ammonium sulfate fractionation and sequential chromatographies on Toyopearl HW65C and DEAE-Toyopearl columns, with an activity recovery of 22.7%. The enzyme is most active at pHs of 9.0 to 9.5. The optimm temperature fot the enzyme is 50-70℃. About 50% of the initial activity remains after incubation at 68℃ and pH 7.5 or 30 min. The isoelectric point of the enzyme is 4.3. Its molecular weight is estimted to be 220,000 Da by gel filtration on FPLC-Hiload Superdex 200 pg and 58,000 Da by SDS-PAGE, suggesting that it is a tetramer. The activity of the enzyme is completely inhibited by PCMB, HgCl_2 and Mn^<2+>. The K_m values of the enzyme for glycerol and ATP are 3.8×10^<-5>M and 1.62×10^<-4>M, resoectively. The N-terminal amino acid sequence of the enzyme is MNQYMLAIDQGTTSSR.

著者

Kabashima T School Of Pharmaceutical Sciences Nagasaki University
Kabashima Tsutomu School Of Pharmaceutical Sciences Nagasaki University
吉田 豊和 岐阜大・工・生命工
Ito Kiyoshi School of Pharmaceutical Sciences, Nagasaki University
Ito K School Of Pharmaceutical Sciences Nagasaki University
YOSHIMOTO TADASHI School of Pharmaceutical Sciences, Nagasaki University
Yoshimoto Tadashi School Of Pharmaceutical Sciences Nagasaki University
Ito K Sapporo Breweries Ltd. Gumma Jpn
Meng Yang School Of Pharmaceutical Sciences Nagasaki University And Toyobo Tsuruga Institute Of Biotechnology
Ito Kiyoshi School Of Pharmaceutical Sciences Nagasaki University
HUANG Hua-Shan School of Pharmaceutical Sciences, Nagasaki University
YOSHIDA TOMOHIRO School of Pharmaceutical Sciences, Nagasaki University, and Toyobo Tsuruga Institute of Biotechnolog
NISHIYA YOSHIAKI School of Pharmaceutical Sciences, Nagasaki University, and Toyobo Tsuruga Institute of Biotechnolog
KAWAMURA YOSHIHISA School of Pharmaceutical Sciences, Nagasaki University, and Toyobo Tsuruga Institute of Biotechnolog
Huang Huashan School Of Pharmaceutical Sciences Nagasaki University
Nishiya Yoshiaki School Of Pharmaceutical Sciences Nagasaki University And Toyobo Tsuruga Institute Of Biotechnology
Kawamura Yoshihisa School Of Pharmaceutical Sciences Nagasaki University And Toyobo Tsuruga Institute Of Biotechnology
Yoshida Tomohiro School Of Pharmaceutical Sciences Nagasaki University And Toyobo Tsuruga Institute Of Biotechnology
Ito K Yamagata Univ. Yonezawa Jpn
Yoshimoto Tadashi School of Pharamaceutical Sciences, Nagasaki University

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