Dinitrogenase with altered substrate specificity results from the use of homocitrate analogs for in vitro synthesis of the iron-molybdenum cofactor

Biochemistry ◽  
1988 ◽  
Vol 27 (10) ◽  
pp. 3647-3652 ◽  
Author(s):  
Timothy R. Hoover ◽  
Juan Imperial ◽  
Jihong Liang ◽  
Paul W. Ludden ◽  
Vinod K. Shah
Keyword(s):  
Substrate Specificity ◽  
Molybdenum Cofactor ◽  
In Vitro Synthesis ◽  
Iron Molybdenum Cofactor

scholarly journals In vitro synthesis of the iron-molybdenum cofactor of nitrogenase.

1986 ◽  
Vol 83 (6) ◽  
pp. 1636-1640 ◽  
Author(s):  
V. K. Shah ◽  
J. Imperial ◽  
R. A. Ugalde ◽  
P. W. Ludden ◽  
W. J. Brill
Keyword(s):  
Molybdenum Cofactor ◽  
In Vitro Synthesis ◽  
Iron Molybdenum Cofactor

scholarly journals In vitro synthesis of the iron molybdenum cofactor of nitrogenase from iron, sulfur, molybdenum, and homocitrate using purified proteins

2007 ◽  
Vol 104 (45) ◽  
pp. 17626-17631 ◽  
Author(s):  
L. Curatti ◽  
J. A. Hernandez ◽  
R. Y. Igarashi ◽  
B. Soboh ◽  
D. Zhao ◽  
...  
Keyword(s):  
Molybdenum Cofactor ◽  
In Vitro Synthesis ◽  
Iron Sulfur ◽  
Iron Molybdenum Cofactor

scholarly journals Requirement of NifX and Other nif Proteins for In Vitro Biosynthesis of the Iron-Molybdenum Cofactor of Nitrogenase

1999 ◽  
Vol 181 (9) ◽  
pp. 2797-2801 ◽  
Author(s):  
Vinod K. Shah ◽  
Priya Rangaraj ◽  
Ranjini Chatterjee ◽  
Ronda M. Allen ◽  
Jon T. Roll ◽  
...  
Keyword(s):  
Azotobacter Vinelandii ◽  
Molybdenum Cofactor ◽  
In Vitro Synthesis ◽  
Mutant Strains ◽  
Dinitrogenase Reductase ◽  
In Vitro Biosynthesis ◽  
Nif Proteins ◽  
Iron Molybdenum Cofactor ◽  
Fold Stimulation

ABSTRACT The iron-molybdenum cofactor (FeMo-co) of nitrogenase contains molybdenum, iron, sulfur, and homocitrate in a ratio of 1:7:9:1. In vitro synthesis of FeMo-co has been established, and the reaction requires an ATP-regenerating system, dithionite, molybdate, homocitrate, and at least NifB-co (the metabolic product of NifB), NifNE, and dinitrogenase reductase (NifH). The typical in vitro FeMo-co synthesis reaction involves mixing extracts from two different mutant strains of Azotobacter vinelandii defective in the biosynthesis of cofactor or an extract of a mutant strain complemented with the purified missing component. Surprisingly, the in vitro synthesis of FeMo-co with only purified components failed to generate significant FeMo-co, suggesting the requirement for one or more other components. Complementation of these assays with extracts of various mutant strains demonstrated that NifX has a role in synthesis of FeMo-co. In vitro synthesis of FeMo-co with purified components is stimulated approximately threefold by purified NifX. Complementation of these assays with extracts of A. vinelandii DJ42.48 (ΔnifENX ΔvnfE) results in a 12- to 15-fold stimulation of in vitro FeMo-co synthesis activity. These data also demonstrate that apart from the NifX some other component(s) is required for the cofactor synthesis. The in vitro synthesis of FeMo-co with purified components has allowed the detection, purification, and identification of an additional component(s) required for the synthesis of cofactor.

Accumulation of factor F395 in nifNE mutants of Klebsiella pneumoniae

1993 ◽  
Vol 39 (3) ◽  
pp. 351-354
Author(s):  
Diana M. Downs ◽  
Paul W. Ludden ◽  
Vinod K. Shah
Keyword(s):  
Nitrogen Fixation ◽  
Klebsiella Pneumoniae ◽  
Small Molecule ◽  
Molybdenum Cofactor ◽  
Gene Products ◽  
In Vitro Synthesis ◽  
Iron Molybdenum Cofactor

The nifNE gene products of Klebsiella pneumoniae are required for the in vivo and in vitro synthesis of the iron-molybdenum cofactor (FeMo-co) of nitrogenase. Derepression of nifNE mutants for nitrogenase resulted in the accumulation of a small molecule, factor F395. Factor F395 is protein associated in vivo. We report here initial spectral characterization of this factor.Key words: FeMo-co, nitrogen fixation, Klebsiella pneumoniae, nifNE.

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