tailieunhanh - Báo cáo Y học: Purification and catalytic properties of a CO-oxidizing:H2-evolving enzyme complex from Carboxydothermus hydrogenoformans

From the membrane fraction of the Gram-positive bacter-ium Carboxydothermus hydrogenoformans,anenzyme complex catalyzing the conversionofCOtoCO2andH2was purified. The enzyme complex showed maximal CO-oxidi-zing:H2 -evolving enzyme activity with 5% CO in the head-space (450 U per mg protein). Higher CO concentrations inhibited the hydrogenase present in the enzyme complex. For maximal activity, the enzyme complex had to be activated by either CO or strong reductants. | Eur. J. Biochem. 269 5712-5721 2002 FEBS 2002 doi Purification and catalytic properties of a CO-oxidizing H2-evolving enzyme complex from Carboxydothermus hydrogenoformans Basem Soboh1 Dietmar Linder2 and Reiner Hedderich1 1 Max-Planck-Institut fur terrestrische Mikrobiologie Karl-von-Frisch-Strafie Marburg Germany 2Biochemisches Institut Fachbereich Humanmedizin Justus-Liebig-Universitat Giessen Germany From the membrane fraction of the Gram-positive bacterium Carboxydothermus hydrogenoformans an enzyme complex catalyzing the conversion of CO to CO2 and H2 was purified. The enzyme complex showed maximal CO-oxidi-zing H2-evolving enzyme activity with 5 CO in the headspace 450 U per mg protein . Higher CO concentrations inhibited the hydrogenase present in the enzyme complex. For maximal activity the enzyme complex had to be activated by either CO or strong reductants. The enzyme complex also catalyzed the CO- or H2-dependent reduction of methylviologen at 5900 and 180 U per mg protein respectively. The complex was found to be composed of six hydrophilic and two hydrophobic polypeptides. The aminoterminal sequences of the six hydrophilic subunits were determined allowing the identification of the encoding genes in the preliminary genome sequence of C. hydrogenofor-mans. From the sequence analysis it was deduced that the enzyme complex is formed by a Ni-containing carbon monoxide dehydrogenase CooS an electron transfer protein containing four 4Fe-4S clusters CooF and a membrane bound NiFe hydrogenase composed of four hydrophilic subunits and two membrane integral subunits. The hydrogenase part of the complex shows high sequence similarity to members of a small group of NiFe hydrogenases with sequence similarity to energy conserving NADH quinone oxidoreductases. The data support a model in which the enzyme complex is composed of two catalytic sites a CO-oxidizing site and a H2-forming site which are connected via a different .