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Induction of nutritional mutants of Micrococcus glutamicus and their amino acid accumulation
Abstract:
Micrococcus glutamicus ATCC 13032, a glutamic acid-producing organism, was treated with 0.2M ethylmethane sulfonate, the auxotrophs isolated showing varied patterns of extracellular amino acids. Eighty auxotrophic strains were obtained, out of which 31 excreted 1.0-4.0 mg threonine per ml and all the auxotrophs required biotin for growth and production of the amino acid. Eleven auxotrophs produced 1.5 to 3.0 mg alanine per ml and these auxotrophs required amino acids for their growth. Other auxotrophs lost their excretion capacity in subsequent fermentation trials. Further mutation of the biotin-requiring auxotroph Micrococcus glutamicus EM with gamma rays resulted in the isolation of 89 auxotrophic strains, out of which 28 excreted threonine (up to 5.0 mg per ml) higher than the parent auxotroph. Exposure to X-rays yielded 97 auxotrophs, out of these 35 producing 1.0-3.0 mg methionine per ml and requiring biotin for growth and production of the amino acid. Other auxotrophs produced alanine (0.5 to 2.0 mg per ml) and threonine (2.0 to 3.3 mg per ml). Irradiation with gamma rays favoured the development of threonine producing auxotrophs while X-rays favoured methionine-producing auxotrophs.
Insights
Mutagenesis of Micrococcus glutamicus generated auxotrophic strains. Gamma rays favored threonine production, while X-rays enhanced methionine production in these amino acid-producing microorganisms.
Area of Science:
- Microbiology
- Biotechnology
- Metabolic Engineering
Background:
- Micrococcus glutamicus is a key microorganism for industrial amino acid production.
- Generating auxotrophic mutants is a strategy to enhance microbial production of specific metabolites.
- Understanding the metabolic pathways and regulatory mechanisms is crucial for strain improvement.
Purpose of the Study:
- To generate and characterize auxotrophic mutants of Micrococcus glutamicus for enhanced amino acid production.
- To investigate the effects of different mutagens (ethylmethane sulfonate, gamma rays, X-rays) on amino acid excretion patterns.
- To identify specific auxotrophic strains with improved threonine, alanine, or methionine production capabilities.
Main Methods:
- Chemical mutagenesis of Micrococcus glutamicus ATCC 13032 using ethylmethane sulfonate.
- Isolation and screening of auxotrophic strains for amino acid production.
- Further mutagenesis of a biotin-requiring auxotroph using gamma rays and X-rays.
- Fermentation trials to quantify extracellular amino acid yields (threonine, alanine, methionine).
Main Results:
- Ethylmethane sulfonate mutagenesis yielded 80 auxotrophs, with 31 excreting threonine (1.0-4.0 mg/ml) and 11 excreting alanine (1.5-3.0 mg/ml).
- Gamma ray irradiation of a biotin-requiring mutant produced 89 auxotrophs, 28 of which excreted higher levels of threonine (up to 5.0 mg/ml).
- X-ray irradiation yielded 97 auxotrophs, with 35 producing methionine (1.0-3.0 mg/ml) and others producing alanine and threonine.
Conclusions:
- Different mutagens differentially affect amino acid production in Micrococcus glutamicus auxotrophs.
- Gamma rays are effective for isolating high-threonine-producing strains, while X-rays favor methionine production.
- Auxotrophic mutants represent a valuable resource for developing improved microbial cell factories for amino acid synthesis.