Abstract

Molybdenum metabolism will be studied in Klebsiella pneumoniae and Azotobacter vinelandii. The biosynthetic pathway of molybdenum into the iron molybdenum cofactor (FeMo-co) of nitrogenase will be investigated. Mutants are available in a number of steps in the pathway, and these will be used to establish steps in the pathway. Intermediates will be purified and characterized with respect to their composition and other properties whenever possible. A long-term goal of the project will be to completely define the pathway of FeMo-co biosynthesis from uptake of molybdate and other molybdenum compounds to the insertion of the cofactor into the FeMo-co--less dinitrogenase. The nifQBNE and V gene products are involved in FeMo-co biosynthesis. These proteins and any other proteins found to be involved in FeMo-co biosynthesis will be purified and characterized. Protein is a limiting factor in human nutrition, and biological nitrogen fixation is the largest source of fixed nitrogen for protein synthesis on earth. Molybdoenzymes are important in human nutrition and in every biological system. The study of molybdenum metabolism and nitrogen fixation will provide information about how biological systems assimilate and process this important trace element.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM035332-01A1
Application #
3287903
Study Section
Microbial Physiology and Genetics Subcommittee 2 (MBC)
Project Start
1986-12-01
Project End
1989-11-30
Budget Start
1986-12-01
Budget End
1987-11-30
Support Year
1
Fiscal Year
1987
Total Cost
Indirect Cost

  Institution

Name
University of Wisconsin Madison
Department
Type
Earth Sciences/Resources
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715

  Publications

Hernandez, Jose A; Phillips, Aaron H; Erbil, W Kaya et al. (2011) A sterile alpha-motif domain in NafY targets apo-NifDK for iron-molybdenum cofactor delivery via a tethered domain. J Biol Chem 286:6321-8
Soboh, Basem; Boyd, Eric S; Zhao, Dehua et al. (2010) Substrate specificity and evolutionary implications of a NifDK enzyme carrying NifB-co at its active site. FEBS Lett 584:1487-92
Hernandez, Jose A; Curatti, Leonardo; Aznar, Constantino P et al. (2008) Metal trafficking for nitrogen fixation: NifQ donates molybdenum to NifEN/NifH for the biosynthesis of the nitrogenase FeMo-cofactor. Proc Natl Acad Sci U S A 105:11679-84
George, Simon J; Igarashi, Robert Y; Piamonteze, Cinthia et al. (2007) Identification of a Mo-Fe-S cluster on NifEN by Mo K-edge extended X-ray absorption fine structure. J Am Chem Soc 129:3060-1
Curatti, Leonardo; Hernandez, Jose A; Igarashi, Robert Y et al. (2007) In vitro synthesis of the iron-molybdenum cofactor of nitrogenase from iron, sulfur, molybdenum, and homocitrate using purified proteins. Proc Natl Acad Sci U S A 104:17626-31
Hernandez, Jose A; Igarashi, Robert Y; Soboh, Basem et al. (2007) NifX and NifEN exchange NifB cofactor and the VK-cluster, a newly isolated intermediate of the iron-molybdenum cofactor biosynthetic pathway. Mol Microbiol 63:177-92
Soboh, Basem; Igarashi, Robert Y; Hernandez, Jose A et al. (2006) Purification of a NifEN protein complex that contains bound molybdenum and a FeMo-Co precursor from an Azotobacter vinelandii DeltanifHDK strain. J Biol Chem 281:36701-9
Dyer, David H; Rubio, Luis M; Thoden, James B et al. (2003) The three-dimensional structure of the core domain of Naf Y from Azotobacter vinelandii determined at 1.8-A resolution. J Biol Chem 278:32150-6
Ruttimann-Johnson, Carmen; Rubio, Luis M; Dean, Dennis R et al. (2003) VnfY is required for full activity of the vanadium-containing dinitrogenase in Azotobacter vinelandii. J Bacteriol 185:2383-6
Rubio, Luis M; Rangaraj, Priya; Homer, Mary J et al. (2002) Cloning and mutational analysis of the gamma gene from Azotobacter vinelandii defines a new family of proteins capable of metallocluster binding and protein stabilization. J Biol Chem 277:14299-305

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