Apo MoFe protein

The biosynthesis of the MoFe protein is extremely complex. Here we will first describe the biosynthesis of FeMoco, then that of the apo-MoFe protein, encoded by the nifD and nifK genes and containing the P clusters, and finally we will summarize what is known about the combination of FeMoco with the apo-MoFe protein to form active MoFe protein. 

There is now some understanding of the processes involved in insertion of FeMoco into the a-ifii polypeptides (see Section IV,C,3). However, it is clear that there are other processes involved in generating apo-MoFe protein capable of being activated. These processes in-... 

The term apo-MoFe protein will be used here to denote the MoFe protein lacking FeMoco, although this protein is not devoid of prosthetic groups since it has bound P clusters. 

The purified preparations of the apo-MoFe proteins from both organisms included a small additional polypeptide of around 20 kDa. This was shown to be the nifY product in K. pneumoniae (94) and a non-nif protein denoted y in A. vinelandii (95). These proteins are apparently essential for effective reaction with FeMoco and are associated with the MoFe protein polypeptide through its interaction with the Fe protein and MgATP (96, 97) (see Section IV,C,3). These observations demonstrate a third role for the Fe protein in generating a form of apo-MoFe protein that is capable of accepting FeMoco. 

An Fe-only nitrogenase has also been isolated from a nifH mutant of Rhodospirillum rubrum and was characterized as an a2/82<% hex-amer containing only iron, no molybdenum or vanadium, with an o 2Fe4S4-containing Fe protein. A factor could be extracted from the FeFe protein into NMF that combined with apo-MoFe protein to form an active enzyme 193). 

Apo MoFe protein, 47 176, 180-181, 182 Apoplastocyanin, structure, 36 384-385 Apoproteins, selenium-substituted, preparation, 38 80-82... 

When FeMoco extracted from MoFe protein purified from a nifV mutant is recombined with apo-MoFe protein, the activated protein has the substrate-reducing characteristics of the nifV enzyme (reduces C2H2 effectively but N2 only poorly). This observation provides the most compelling evidence that FeMoco is, or forms part of, the active site of nitrogenase. Site-directed mutagenesis has implicated one of the conserved Cys residues of the a subunit Cys 275 in binding FeMoco, and also His 196 and Gin 192 (see Refs. 17 and 38 for discussion). 

Finally, the FeMoeo is transferred to nifDK (apo-MoFe protein) to form the holo-MoFe protein with the help of ATP and several proteins nifH, 7, and GroEL. The X-ray crystal structure of nifDK shows a positively charged channel through which FeMoeo could enter. " The mf system for synthesis of the FeVco is thought to be similar to the nif system, with added assistance in the last step by vnfG, a third subunit of VFe nitrogenase that is necessary for activity. 

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