CHARACTERIZATION OF THE ACTIVE-SITE OF A HYDROGEN SENSOR FROM ALCALIGENES-EUTROPHUS

A third hydrogenase was recently identified in the proteobacterium Alc aligenes eutrophus as a constituent of a novel H-2-sensing multicompon ent regulatory system. This regulatory hydrogenase (RH) has been overe xpressed in cells deficient in both the NAD(+)-reducing [NiFe]-hydroge nase and the membrane-bound [NiFe]-hydrogenase. EPR, FTIR and activity studies of membrane-free extracts revealed that the RH has an active site much like that of standard [NiFe]-hydrogenases, i.e. a Ni-Fe site with two CN- groups and one CO molecule. Its catalytic power is low, but the RH is always active, insensitive to oxygen, and occurs in only two redox states. (C) 1998 Federation of European Biochemical Societi es.