REACTIVITY OF THE N-TERMINAL CYSTEINE RESIDUES IN ACTIVE AND INACTIVEFORMS OF FNR, AN O-2-RESPONSIVE, FE CONTAINING TRANSCRIPTIONAL REGULATOR OF ESCHERICHIA-COLI

FNR, the O-2-responsive gene regulator of anaerobic respiratory genes in Escherichia coli, contains an N-terminal cluster of four cysteine r esidues (Cys16-X(3)-Cys20-X(2)-Cys23-X(5)-Cys29), three of which are t hought to be involved in the binding of an iron cofactor. The accessib ility of the cysteine residues for iodoacetate is known to increase up on switch from the active (anaerobic)to the inactive (aerobic or metal depleted) state. It was analyzed which residues become accessible und er either condition. Up to four modified forms, FNR-I, FNR-II, FNR-III , and FNR-IV, containing approximately 1, 2, 3.5, and 5 carboxymethyl groups, were obtained either by reaction in vivo and in vitro under co nditions of aerobiosis, anaerobiosis, or iron limitation. By N-termina l sequencing, the carboxymethylated cysteine residues were identified. The amount of label in each of the four cysteine residues increased r ather uniformly and gradually from FNR-I to FNR-IV irrespective of the condition of labeling; only Cys16 was preferentially labeled to some extent. It is concluded that the four essential cysteine residues chan ge their accessibility in a similar way in the switch from active to i nactive (aerobic or metal depleted) FNR, without specific differences in their reaction or function. Potential modes of Fe-binding by the cy steine residues are discussed. In addition, a different type of intera ction of Fe(II) with FNR is described. The interaction occurred also i n FNR carboxymethylated at approximately three cysteine residues.