Microbial synthesis of 2H-labelled L-phenylalanine with different levels of isotopic enrichment by a facultative methylotrophic bacterium Brevibacterium methylicum with RuMP assimilation of carbon

Autor: O. V. Mosin, I. Ignatov, V. I. Shvets, D. A. Skladnev
Rok vydání: 2013
Předmět:
Zdroj: Biochemistry (Moscow) Supplement Series B: Biomedical Chemistry. 7:247-258
ISSN: 1990-7516
1990-7508
DOI: 10.1134/s1990750813030098
Popis: Using the L-phenylalanine secreting strain of Gram-negative aerobic facultative methylotrophic bacteria Brevibacterium methylicum, assimilating methanol via the ribulose-5-monophosphate (RuMP) cycle of carbon assimilation, as an example, we have continued studies on the use of methylotrophic bacteria for the preparative microbial synthesis of amino acids labeled with stable isotopes, including deuterium (2H), suitable for biomedical applications and clinical diagnostics. Here we demonstrate the data on adaptation of the methylotrophic bacterium B. methylicum to the maximal concentration of deuterium in the growth medium with 98% (v/v) 2H2O and 2% (v/v) [2H]MeOH, and biosynthesis of deuterium labeled L-phenylalanine with different levels of isotopic enrichment. The strain was adapted to 2H2O by means of plating of initial cells on solid (2% agarose) minimal growth media M9 with an increasing gradient of 2H2O concentration from 0, 24.5, 49.0, 73.5 up to 98% (v/v) 2H2O and subsequent selection of individual colonies stable to the action of 2H2O, which were capable to produce L-phenylalanine. L-phenylalanine was extracted from the growth medium with isopropanol followed by subsequent crystallization in ethanol (output 0.65 g/L). Using the developed method of microbial synthesis it is possible to obtain deuterated L-phenylalanine with different levels of isotopic enrichment, depending on concentration of 2H2O in growth media, from 17% (the growth medium with 24.5% (v/v) 2H2O) right up to 75% (the growth medium with 98% (v/v) 2H2O) of deuterium as evidenced by results of the electron impact (EI) mass-spectrometry analysis of methyl ethers of N-dimethylamino(naphthalene)-5-sulfonyl chloride (dansyl) phenylalanine isolated from growth media under different experimental conditions.
Databáze: OpenAIRE
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