Open AccessNAD‐Dependent Formate Dehydrogenase from Methylotrophic Bacterium, Strain 1
Author(s) -
EGOROV Aleksey M.,
AVILOVA Tatyana V.,
DIKOV Michael M.,
POPOV Vladimir O.,
RODIONOV Yuriy V.,
BEREZIN Ilia V.
Publication year - 1979
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1979.tb13289.x
Subject(s) - formate dehydrogenase , formate , chemistry , nad+ kinase , ammonium formate , chromatography , dehydrogenase , biochemistry , polyacrylamide gel electrophoresis , gel electrophoresis , enzyme , size exclusion chromatography , catalysis , mass spectrometry
1. NAD‐dependent formate dehydrogenase was isolated from gram‐negative methylotrophic bacteria, strain 1, grown on methanol. The purification procedure involved ammonium sulfate fractionation, ion‐exchange chromatography and preparative isotachophoresis or gel filtration; it resulted in a yield of 40 %. 2. The final enzyme preparations were homogeneous as judged by sedimentation in an ultra‐centrifuge. Formate dehydrogenase purified in the presence of EDTA reveals two bands on electrophoresis in polyacrylamide gel both after protein and activity staining. Two components are transformed into a single one after prolonged storage in the presence of 2‐mercaptoethanol. 3. Formate dehydrogenase is a dimer composed of identical or very similar subunits. The molecular weight of the enzyme is about 80000. 4. Amino acid composition and some other physico‐chemical properties of the enzyme were studied. 5. Formate dehydrogenase is specific for formate and NAD as electron acceptor. The Michaelis constant was 0.11 mM for NAD and 15 mM for formate (pH 7.0, 37°C). 6. Formate dehydrogenase was rapidly inactivated in the absence of –SH compounds. The enzyme retained full activity upon storage at ambient temperature in solution for half a year in the presence of 2‐mercaptoethanol or EDTA.