The element nitrogen (N) is essential to all living organisms, and the availability of fixed forms of nitrogen is often a limiting factor in food production. Biological nitrogen fixation, which occurs in a diverse group of microbes, represents the single largest input of fixed nitrogen in the reductive phase of the global nitrogen cycle. A molecular understanding of the enzyme responsible for biological nitrogen fixation, nitrogenase, could contribute to enhanced food production worldwide, and thus to the overall health of the human population. In addition, as a representative of the diverse family of metalloproteins and nucleotide-dependent enzymes, understanding the mechanism of nitrogenase will contribute to a broader understanding of these important enzymes. Our goal is to use a comprehensive approach to address some of the significant remaining questions about the nitrogenase mechanism. The related mechanistic issues that will be examined include the following: (i) understanding how MgATP hydrolysis is coupled to substrate reduction; (ii) defining how electron transfer through nitrogenase is controlled; (iii) understanding the roles of the associated metal clusters in this process; and (iv) defining how substrates access and products exit the active site. These goals will be accomplished by use of a wide array of methods including genetic selections, site-directed mutagenesis, purification of nitrogenase proteins with individual and multiple amino acid substitutions, steady state and pre-steady-state kinetics, electrochemistry, and spectroscopic methods including EPR, ENDOR, and X-ray crystallography.
| Hoffman, Brian M; Lukoyanov, Dmitriy; Yang, Zhi-Yong et al. (2014) Mechanism of nitrogen fixation by nitrogenase: the next stage. Chem Rev 114:4041-62 |
| Lukoyanov, Dmitriy; Yang, Zhi-Yong; Duval, Simon et al. (2014) A confirmation of the quench-cryoannealing relaxation protocol for identifying reduction states of freeze-trapped nitrogenase intermediates. Inorg Chem 53:3688-93 |
| Duval, Simon; Danyal, Karamatullah; Shaw, Sudipta et al. (2013) Electron transfer precedes ATP hydrolysis during nitrogenase catalysis. Proc Natl Acad Sci U S A 110:16414-9 |
| Hoffman, Brian M; Lukoyanov, Dmitriy; Dean, Dennis R et al. (2013) Nitrogenase: a draft mechanism. Acc Chem Res 46:587-95 |
| Yang, Zhi-Yong; Khadka, Nimesh; Lukoyanov, Dmitriy et al. (2013) On reversible H2 loss upon N2 binding to FeMo-cofactor of nitrogenase. Proc Natl Acad Sci U S A 110:16327-32 |
| Seefeldt, Lance C; Yang, Zhi-Yong; Duval, Simon et al. (2013) Nitrogenase reduction of carbon-containing compounds. Biochim Biophys Acta 1827:1102-11 |
| Mayweather, Diana; Danyal, Karamatullah; Dean, Dennis R et al. (2012) Correction to temperature invariance of the nitrogenase electron transfer mechanism. Biochemistry 51:9027 |
| Seefeldt, Lance C; Hoffman, Brian M; Dean, Dennis R (2012) Electron transfer in nitrogenase catalysis. Curr Opin Chem Biol 16:19-25 |
| George, Simon J; Barney, Brett M; Mitra, Devrani et al. (2012) EXAFS and NRVS reveal a conformational distortion of the FeMo-cofactor in the MoFe nitrogenase propargyl alcohol complex. J Inorg Biochem 112:85-92 |
| Yang, Zhi-Yong; Moure, Vivian R; Dean, Dennis R et al. (2012) Carbon dioxide reduction to methane and coupling with acetylene to form propylene catalyzed by remodeled nitrogenase. Proc Natl Acad Sci U S A 109:19644-8 |
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