Purification and characterization of the vnf-encoded apodinitrogenase from Azotobacter vinelandii.
The vnf-encoded apodinitrogenase (apodinitrogenase 2) has been purified from Azotobacter vinelandii strain CA117.30 (DeltanifKDB), and is an alpha2beta2delta2 hexamer. Apodinitrogenase 2 can be activated in vitro by the addition of the iron-vanadium cofactor (FeV-co) to form holodinitrogenase 2, which functions in C2H2, H+, and N2 reduction. Under certain conditions, the alpha2beta2delta2 hexamer dissociates to yield the free delta subunit (the VNFG protein) and a form of apodinitrogenase 2 that exhibits no C2H2, H+, or N2 reduction activities in the in vitro FeV-co activation assay; however, these activities can be restored upon addition of VNFG to the FeV-co activation assay system. No other vnf-, nif-, or non-nif-encoded proteins were able to replace the function of VNFG in the in vitro processing of alpha2beta2 apodinitrogenase 2 (in the presence of FeV-co) to a form capable of substrate reduction. Apodinitrogenase 2 is also activable in vitro by the iron-molybdenum cofactor to form a hybrid enzyme with unique properties, most notably the inability to reduce N2 and insensitivity to CO inhibition of C2H2 reduction.
Chatterjee
R
R
Department of Biochemistry, College of Agricultural and Life Sciences, University of Wisconsin-Madison, 53706, USA.
Allen
R M
RM
Ludden
P W
PW
Shah
V K
VK
D016948:Azotobacter vinelandii / Q000201:enzymology*; D004789:Enzyme Activation; D009591:Nitrogenase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity
D009591:Nitrogenase