Characterisation of a protease from E. coli involved in hydrogenase maturation.

The large subunits of nickel-containing hydrogenases are synthesised in a precursor form which, after nickel incorporation, is processed by proteolytic cleavage at the C-terminal end. The protease involved in processing of HycE. the large subunit of hydrogenase 3 from Escherichia coli, was purified by three chromatographic steps to apparent homogeneity. Its gene was identified by using a hybridisation probe generated by PCR with oligonucleotide primers the sequence of which was derived from the N-terminal and internal amino acid sequences. Determination of tile nucleotide sequence showed that the gene is located distally and as a hitherto uncharacterised gene within the hyc operon, coding for hydrogenase 3 components. It was designated hycI. The HycI protease has a molecular mass of 17 k Da and is a monomer. Its cleavage reaction is not inhibited by conventional inhibitors of serine and metalloproteases, which correlates with the fact that the sequence does not contain signal are motifs characteristic of serine-, metallo-, cysteine- or acid proteases. Homologous genes are present in other transcriptional units coding for hydrogenases. [ABSTRACT FROM AUTHOR]