Nitrogen is an element that is essential for all life processes and it is found abundantly in the Earth's atmosphere in the form of N2 gas. However, N2 is not metabolized by most organisms and consequently, cellular nitrogen is usually obtained as ammonia, nitrate, or as part of an organic molecule. These combined sources of nitrogen are relatively scarce and represent the most limiting nutrient in Man's effort to feed the World population. Because organic nitrogen is incompletely recycled in the global ecosystem and available ammonia and nitrate are continually metabolized to N2 through nitrification and denitrification, the biological reduction of N2 occupies a pivotal position in the nitrogen cycle. This process, called nitrogen fixation, is performed only by certain prokaryotic microorganisms and is catalyzed by the enzyme nitrogenase. The agronomic significance of symbiotic microorganisms that fix N2 and deliver it to their plant hosts is well established and there are real prospects that the economy of nitrogen fixation can be improved through the genetic manipulation of N2-fixing species. Such improvements are dependent upon a fundamental understanding of the biochemical action and genetic regulation of the nitrogen fixation should prove invaluable in duplicating the activity of nitrogenase in chemical systems, which might lead to viable complementary commercial processes. The proposed research is centered on the iron-molybdenum cofactor, the putative N2-binding site of nitrogenase, to determine how nitrogenase is organized to effect biological nitrogen fixation. Experiments include: modification, purification, and/or crystallization to determine its chemical composition, size and structure; electrochemical, chemical and spectroscopic studies to elucidate its redox properties and potential for catalysis; characterization of potentially altered cofactors from mutant strains; how and where the cofactor is bound to the polypeptides; and how this interaction modifies its structure and functioning. The resulting information should aid both in the understanding of the catalytic mechanism and also in generating targets for beneficial modifications of the enzyme, such that a significant health and nutritional benefit might accrue in the future.
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