Document Type

Thesis - Open Access

Award Date


Degree Name

Master of Science (MS)



First Advisor

Robert M. Pengra


Biological nitrogen fixation, the mechanism of the enzymatic conversion of the inert nitrogen gas into amino acids in the cytoplasm of the microorganism has puzzled many scientists for decades. The recent discovery of some free-living nitrogen-fixing bacteria has encouraged the investigation of this mechanism. A big step taken in the rapid elucidation of this mechanism was the first reproducible cell-free nitrogen-fixing system from Clostridium pasteurianum successfully obtained by Carnahan et al. (9) in 1960. In the following decade, reproducible cell-free systems were demonstrated with the strict aerobe, Azotobacter vinelandii (4), facultative anaerobic Bacillus polymyxa (16) and Klebsiella pneumoniae (31) and photosynthetic bacteria, Rhodospirillum rubrum (46) and Chromatium (1). The first major advance produced as the result of development of the cell-free system was the understanding of the requirement for an energy source and an electron donor for nitrogen fixation. The energy requirement in all cell-free systems is satisfied by the biological energy source, ATP. The most effective reductant found to date for all the cell-free systems is the artificial reductant, dithionite (5, 6, 16, 31). The natural electron donor and the electron pathway were subsequently studied in some bacteria. Most extensively studied were the strict anaerobe C. pasteurianum and the strict aerobe A. vinelandii. Several intermediary metabolites, such as pyruvate, α-ketobutyrate (9), formate and malate (34) have been found to function well as reductants for the nitrogenase of C. pasteurianum while none of these or other natural substrates are effective with extracts of A. vinelandii (20). Thus, the survey of the natural electron donor in nitrogen fixation by the facultative bacterium K. pneumoniae may cast light on the mystery of the mechanism of nitrogen fixation by these bacteria. The object of the present investigation was to serve this purpose.

Library of Congress Subject Headings

Nitrogen -- Fixation
Nitrogen -- fixing microorganisms
Enterobacter aerogenes



Number of Pages



South Dakota State University

Included in

Biology Commons