Characterization by X-ray absorption spectroscopy of the biosynthesis of the cofactors of nitrogenase
Abstract/Contents
- Abstract
- A group of catalytic enzymes known as nitrogenases is able to perform the chemical feat of reducing atmospheric N2 to NH3, a bioavailable nitrogen source for the assembly of biological macromolecules, due largely to the complex electronic and geometric structures of their associated transition metal cofactors. The nitrogenase of Azotobacter vinelandii consists of a two-component system, the MoFe protein, a substrate fixing enzyme, and its specific reductase, the Fe protein. Together the two form a catalytic complex under turnover. An electron transport sequence is formed originating with the [Fe4S4] cofactor of the Fe protein proceeding to the P-cluster (Fe8S7) of the MoFe protein and ending with the iron-molybdenum cofactor (FeMoco), a MoFe7S9C cluster also of the MoFe protein, where substrate reduction occurs. The biosynthesis of these cofactors is an exceedingly involved process still not fully understood, but which serves as a model for cofactor assembly in other biological systems and a template for generation of functional synthetic analogues. Using a carefully constructed series of experiments involving genetic manipulation, reconstitution assays, and structural determination by X-ray Absorption Spectroscopy, a sequence of critical and essential events in cofactor synthesis, requisite gene products, and previously unidentified intermediates have been characterized. Despite a high degree of sequence homology, remarkable differences were detected in the structure of the [Fe4S4] cofactor of alternative nitrogenase Fe proteins. The stepwise fusion of P-cluster on [delta]nifH and [delta]nifB[delta]nifZ MoFe protein was observed to involve an iterative mechanism and the involvement of a new gene product was confirmed. An all iron precursor bound to the NifEN scaffold was determined to have a novel structure and suggest a new mechanism for insertion of heterometal. The solution potential and protein channel were found to have important roles in the maturation of a molybdenum-containing FeMoco precursor. Finally, the structural differences in the iron core of FeMoco and FeVco (a vanadium containing analogous cofactor) were correlated with the differences in substrate reducing profile. Together, these results help to unlock the puzzle of nitrogenase structure and function, a function essential for all life on Earth.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2012 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Blank, Michael Aaron |
|---|---|
| Associated with | Stanford University, Department of Chemistry |
| Primary advisor | Hedman, B. (Britt), 1949- |
| Primary advisor | Hodgson, Keith |
| Thesis advisor | Hedman, B. (Britt), 1949- |
| Thesis advisor | Hodgson, Keith |
| Thesis advisor | Solomon, Edward I |
| Advisor | Solomon, Edward I |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | Michael Aaron Blank. |
|---|---|
| Note | Submitted to the Department of Chemistry. |
| Thesis | Thesis (Ph.D.)--Stanford University, 2012. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2012 by Michael Aaron Blank
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