Comparison of the iron proteins from the nitrogen fixation complexes of Azotobacter vinelandii, Clostridium pasteurianum , and Klebsiella pneumoniae

Abstract

The MW, amino acid compositions, amino- and carboxyl-terminal sequences, and ion-exchange peptide maps of the cysteine-containing tryptic peptides were determined for the Fe proteins from the N2 fixation complexes of A. vinelandii (Av2) and K. pneumoniae (Kp2). Results are compared to the known amino acid sequence of the Fe protein from C. pasteurianum (Cp2). Previous studies showed the Fe proteins to have similar enzymatic functions and spectroscopic properties. DNA coding for the Fe protein from many different species cross-hybridize. The protein structures are similar, yet they have significant differences. The amino-terminal sequences of Av2 and Kp2 are extended, compared to the amino-terminal methionine of Cp2 and may indicate a different initiation site in these proteins. The amino-terminal sequences for Av2 and Kp2 are more homologous with each other than either of these are with Cp2. The carboxyl-terminal sequences are extended in Av2 (14 residues) and Kp2 (.apprxeq. 30 residues) compared to Cp2. The amino- and carboxyl-terminal sequences establish that either the structural gene sizes are different in the 3 organisms or extensive post-translational modification must occur in some species. Because cysteinyl residues are involved at the active site of the Fe protein, a sensitive peptide mapping technique was used to compare cysteinyl peptides of the Fe protein from the 3 species. Av2 and Kp2 have a redistribution of cysteinyl residues when compared to Cp2. Three important differences in the cysteine distributions were found: residue 4 is valine and residue 148 is alanine in Cp2, but cysteinyl residues occupy these positions in Av2; residue 231 is cysteine in Cp2 but alanine in Av2. The peptide mapping technique provides a method for the investigation of selective chemical modification of cysteinyl residues.