Properties of Glucose-Dehydrogenase-Poly(Ethylene Glycol)-NAD Conjugate as an NADH-Regeneration Unit in Enzyme Reactors
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概要
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Glucose-dehydrogenase-poly(ethylene glycol)-NAD conjugate (GlcDH-PEG-NAD) was prepared and its kinetic properties as an NADH-regeneration unit were investigated. The conjugate has about two molecules of active and covalently linked NAD per tetramer. The specific activity of the enzyme moiety of the conjugate in the presence of exogenous NAD is about 77% of that of the native enzyme, and this decrease is mainly due to the decrease in the V_<max> value. The conjugate has the same pH-stability profile as the native enzyme and an internal activity of 0.26s^<-1> (as a monomer); its NAD moiety has similar coenzyme acitivity to poly(ethylene glycol)-bound NAD. These results indicate that GlcDH-PEG-NAD can be used as an NADH-regeneration unit for many dehydrogenase reactions. The coupled reaction of GlcDH-PEG-NAD and lactate dehydrogenase was then studied. The specific activity of the conjugate is 1.1s^<-1> (as a tetramer), the recycling rate of the active NAD moiety is 0.54 s^<-1>, and the apparent K_m value for glucose is 24 mM. Kinetically, lactate dehydrogenase behaves like a substrate with an apparent K_m value of 1.8 units・ml^<-1> in this coupled reaction system with low coenzyme concentration. L-Lactate was continuously produced from pyruvate in a reactor with a PM10 ultrafiltration membrane, and containing GlcDH-PEG-NAD and lactate dehydrogenase. GlcDH-PEG-NAD proved to be applicable in continuous enzyme reactors as an NADH-regeneration unit with a large molecular size.
- 公益社団法人日本生物工学会の論文
- 1988-06-25
著者
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Nakamura A
Department Of Applied Physics Nagoya University
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NAKAMURA Akio
Department of Material Science, Japan Atomic Energy Research Institute
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Urabe I
Osaka Univ. Osaka Jpn
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Urabe Itaru
Department Of Biotechnology Faculty Of Engineering Osaka University
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Urabe Itaru
Dept. Biotech. Grad. Sch. Eng. Osaka Univ.
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Okada H
Kansai Univ. Osaka Jpn
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Okada H
Department Of Fermentation Technology Faculty Of Engineering Osaka University
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Okada H
Nagaoka Univ. Technol. Niigata Jpn
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Okada H
Nagoya City Univ. School Of Medicine Nagoya Jpn
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Okada H
Department Of Fermentation Technology Facutly Of Enginering Osaka Univeristy
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Okada Hirosuke
Deparment Of Fermentation Technology Osaka University
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Urabe Itaru
Department Of Biotechnology Osaka University
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MINAMI HIROKAZU
Department of Fermentation Technology, Faculty of Engineering, Osaka University
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Nakamura Akio
Department Of Information And Computer Sciences Saitama University
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Minami Hirokazu
Department Of Fermentation Technology Faculty Of Engineering Osaka University
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Minami Hirokazu
Department Of Chemistry Faculty Of Engineering Science Osaka University
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