Purification and characterization of two forms of hydrogenase isoenzyme 1 from Escherichia coli.
A hydrogenase associated with dihydrogen uptake (HUP hydrogenase) was purified from an Escherichia coli mutant (strain SE1100) defective in utilization of molybdate and thus fermentative dihydrogen production. This protein had two subunits with apparent molecular weights of 59,000 and 28,000 (form 1). An immunologically cross-reactive hydrogenase was also purified from E. coli K10 grown in glucose-minimal medium and harvested at the mid-exponential phase of growth. Upon purification to homogeneity, this hydrogenase contained only one subunit with an apparent molecular weight of 59,000 (form 2). The two forms of the HUP hydrogenase exhibited similar kinetic characteristics. The electrophoretic properties of the enzyme and its response to pH suggest that this HUP hydrogenase is the HYD1 isoenzyme. The HYD1 isoenzyme was the only hydrogenase detectable during the stationary phase of growth in E. coli grown in Mo-deficient medium.
Department of Microbiology and Cell Science, University of Florida, Gainesville 32611.
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K
Francis
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Patel
J C
JC
Wendt
K T
KT
Shanmugam
D002458:Cell Fractionation / Q000379:methods; D002462:Cell Membrane / Q000201:enzymology; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology*; D007202:Indicators and Reagents; D007527:Isoenzymes / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D010088:Oxidoreductases / Q000302:isolation & purification* / Q000378:metabolism
D007202:Indicators and Reagents; D007527:Isoenzymes; D046911:Macromolecular Substances; D010088:Oxidoreductases; C021376:hydrogen dehydrogenase