NICKEL INCORPORATION INTO HYDROGENASE-3 FROM ESCHERICHIA-COLI REQUIRES THE PRECURSOR FORM OF THE LARGE SUBUNIT

A mutant derivative of hycE, the gene for the large subunit of hydroge nase 3 from Escherichia coli, was constructed that lacks the 3'-termin al part encoding the C-terminal portion of the HycE polypeptide, which is proteolytically removed during maturation of the hydrogenase. The truncated gene was transferred to the in situ position on the chromoso me. Although the mutant possessed HycE in its ''mature'' form, it was devoid of hydrogenase 3 activity. The activity was not restored by hig h nickel concentrations in the medium. The mutated HycE was not associ ated with detectable radioactivity when the strain was grown in the pr esence of Ni-63(2+). These results indicate that the C-terminal extens ion in the precursor form of the large subunit keeps the protein in a conformation required for the coordination of the metal.