Studies on the improvement of lysine production in the genera Brevibacterium and Corynebacterium
Master Thesis
1983
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University of Cape Town
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Abstract
A programme was undertaken to obtain high lysine producing bacteria by mutation of selected wild type strains. Overproduction of glutamic acid under suitable physiological conditions, viz biotin limitation, was chosen as a good indication of the potential of wild type bacteria for improvement in lysine production by mutation. Brevibacterium lactofermentum ATCC 13869 was found to produce the highest amount of glutamic acid under the conditions used. Homoserine and leucine auxotrophic mutants were obtained from this organism and tested for ability to produce lysine. The combination of homnserine and leucine auxotrophy was found to be most effective in overcoming several of the control mechanisms present in the lysine biosynthetic pathway. Lysine production was increased approximately forty fold over the wild type B. lactofermentum. Lysine analogue resistant strains were obtained by further mutation, and lysine production was increased by 20%. The activity and properties of aspartate kinase, a key enzyme in biosynthesis and control of lysine production, was determined to elucidate the nature of the anafogue resistance. Although resistance to feed-back control by lysine and threonine was not responsible for the improvement in lysine production, a considerably higher enzyme activity was found. As a result of the enzyme study a possible novel regulatory system in the lysine biosynthetic pathway of B.lactofermentum ATCC 13869, and the mutants derived from it, was indicated. Environmental optimization studies were undertaken on potentially suitable mutants in order to increase lysine production still further. Fermentation media were improved and a series of fermentations were conducted under precisely controlled conditions in 12 and 20 litre laboratory scale fermenters. The most successful attempt incorporated incremetal feeding of yeast extract and glucose to an S-(2-aminoethyl)-L-cysteine resistant double auxotrophic mutant. A yield of 32 mg/mQ L-lysine.HCI was obtained after 73 hours.
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Bibliography: pages 122-135.
Reference:
Louw, M. 1983. Studies on the improvement of lysine production in the genera Brevibacterium and Corynebacterium. University of Cape Town.