Pilot scale production and isolation of recombinant NAD+- and NADP+-specific formate dehydrogenases.
The expression of the recombinant wild-type NAD+- and mutant NADP+-dependent formate dehydrogenases (EC 1.2.1.2., FDH) from the methanol-utilizing bacterium Pseudomonas sp. 101 in Escherichia coli cells has been improved to produce active and soluble enzyme up to the level of 50% of total soluble proteins. The cultivation process for E. coli/pFDH8a and E. coli/pFDH8aNP cells was optimized and scaled up to a volume of 100 L. A downstream purification process has been developed to produce technical grade NAD+- and NADP+-specific formate dehydrogenases in pilot scale, utilizing extraction in aqueous two-phase systems.
Department of Chemical Enzymology, Faculty of Chemistry, M. V. Lomonosov Moscow State University, 119899 Moscow, Russian Federation. tishkov@ibt022.ibt.kfa-juelich.de
V I
VI
Tishkov
A G
AG
Galkin
V V
VV
Fedorchuk
P A
PA
Savitsky
A M
AM
Rojkova
H
H
Gieren
M R
MR
Kula
D018533:Biomass; D004926:Escherichia coli / Q000201:enzymology; D005285:Fermentation; D005560:Formate Dehydrogenases / Q000737:chemistry* / Q000302:isolation & purification*; D016297:Mutagenesis, Site-Directed; D011549:Pseudomonas / Q000201:enzymology; D011994:Recombinant Proteins / Q000302:isolation & purification; D013997:Time Factors
D011994:Recombinant Proteins; C037212:formate dehydrogenase (NADP+); D005560:Formate Dehydrogenases