Editor: Sylvie Rimsky
Overexpression and biochemical characterization of soluble pyridine nucleotide transhydrogenase from Escherichia coli
Article first published online: 6 MAY 2011
© 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved
FEMS Microbiology Letters
Volume 320, Issue 1, pages 9–14, July 2011
How to Cite
Cao, Z., Song, P., Xu, Q., Su, R. and Zhu, G. (2011), Overexpression and biochemical characterization of soluble pyridine nucleotide transhydrogenase from Escherichia coli. FEMS Microbiology Letters, 320: 9–14. doi: 10.1111/j.1574-6968.2011.02287.x
- Issue published online: 1 JUN 2011
- Article first published online: 6 MAY 2011
- Accepted manuscript online: 16 APR 2011 08:19AM EST
- Received 7 December 2010; revised 9 February 2011; accepted 4 April 2011., Final version published online 6 May 2011.
- soluble pyridine nucleotide transhydrogenase;
- Escherichia coli
The soluble pyridine nucleotide transhydrogenase (STH) is an energy-independent flavoprotein that directly catalyzes hydride transfer between NAD(H) and NADP(H) to maintain homeostasis of these two redox cofactors. The sth gene in Escherichia coli was cloned and expressed as a fused protein (EcSTH). The purified EcSTH displayed maximal activity at 35 °C, pH 7.5. Heat-inactivation studies showed that EcSTH retains 50% activity after 5 h at 50 °C. The enzyme was stable at 4 °C for 25 days. The apparent Km values of EcSTH were 68.29 μM for NADPH and 133.2 μM for thio-NAD+. The kcat/Km ratios showed that EcSTH had a 1.25-fold preference for NADPH over thio-NAD+. Product inhibition studies showed that EcSTH activity was strongly inhibited by excess NADPH, but not by thio-NAD+. EcSTH activity was enhanced by 2 mM adenine nucleotide and inhibited by divalent metal ions: Mn2+, Co2+, Zn2+, Ni2+ and Cu2+. However, after preincubation for 30 min, most divalent metal ions had little effect on EcSTH activity, except Zn2+, Ni2+ and Cu2+. The enzymatic analysis could provide the important basic knowledge for EcSTH utilizations.