L-lysine production at 50 degrees C by mutants of a newly isolated and characterized methylotrophic Bacillus sp

Abstract

The amino acid L-lysine was produced from homoserine auxotrophic and S-(2-aminoethyl)-L-cysteineresistant mutants of a newly isolated gram-positive methylotrophic bacterium, capable of growth on methanol at 60°C. The temperature optimum for growth was between 50 and 53°C. These aerobic, gram-positive, endospore-forming, rod-shaped bacteria required biotin and vitamin B12 for growth. Extracts of the bacteria grown on methanol lacked hydroxypyruvate reductase and contained hexulose 6-phosphate synthase activity. Therefore, these bacteria were considered to be type I methylotrophic bacteria of the genus Bacillus. Fed-batch fermentations resulted in cell densities of 50 g of cell dry weight per liter. Biomass yields on carbon, nitrogen, phosphate, and sulfate were determined. Generation of homoserine auxotrophic and amino acid analogresistant mutants resulted in L-lysine concentrations of nearly 20 g/liter in fed-batch fermentations.

Proteomic analysis of the thermophilic methylotroph Bacillus methanolicusMGA3

“…B. methanolicus MGA3 strain was used throughout the study and was routinely cultivated in minimal salt vitamin (MSV) medium modified after [6] To complete the MSV medium, vitamin stock solution was added prior to use, resulting in the following final concentrations: thiamine hydrochloride, 100 g/L; riboflavin, 100 g/L; pyridoxine hydrochloride, 100 g/L; pantothenate, 100 g/L; nicotinamide, 100 g/L; p-aminobenzoic acid, 20 g/L; folic acid, 10 g/L; cobalamine, 10 g/L; lipoic acid, 10 g/L. All chemicals were obtained from Sigma-Aldrich, Buchs, Switzerland.…”

“…Methylotrophic bacteria utilize reduced one-carbon (C1) sources such as methanol as their sole carbon and energy source [1]. They have in common an initial oxidation step converting methanol to formaldehyde, which is subsequently used for the production of energy and biomass via various cells overproducing L-lysine have been obtained [6]. In addition, transformation methods and expression plasmids for metabolic engineering have been described for this organism [8][9][10].…”

Expression of recombinant green fluorescent protein in Bacillus methanolicus

“…Studies with B. methanolicus to date have principally involved elucidation of the pathway for methanol utilization,3–5 and evaluations of the roles of native enzymes in regulating the production of lysine and glutamate 6–9. Fermentation experiments in previous studies10 have shown that optimal growth conditions include methanol controlled at 150 mM,3 growth temperature of 50°C, pH of 6.8 and media containing biotin, vitamin B 12 , and a small amount of yeast extract in addition to salts 11. The methanol concentration of 150 mM was shown to maximize the utilization of the assimilatory RuMP pathway leading to cell growth and metabolism as opposed to rapid dissimilation of methanol to CO 2 from formaldehyde that occurs in the presence of high methanol concentrations 3.…”

Composition analysis and minimal treatments to solubilize polysaccharides from the brown seaweed Laminaria digitata for microbial growth of thermophiles

“…The Gram-positive bacterium Bacillus methanolicus is a moderate thermophile. It is an aerobic facultative methylotroph and can utilize a few single carbon sources, including mannitol, for growth (Schendel et al 1990;Arfman et al 1992;López et al 2019). Growth of B. methanolicus in 2% NaCl and adaptation to rapid growth in seawater-based media has been described previously (Arfman et al 1992;Komives et al 2005).…”