Microbiological Synthesis of 2H-Labeled Phenylalanine, Alanine, Valine, and Leucine/Isoleucine with Different Degrees of Deuterium Enrichment by the Gram-Positive Facultative Methylotrophic Bacterium Вrevibacterium Methylicum
¹Moscow State University of Applied Biotechnology, Moscow, Russian Federation
²Scientific Research Center of Medical Biophysics (SRCMB), Sofia, Bulgaria
* Corresponding author: Dr. Oleg V. Mosin, PhD. Scientist employee Moscow State University of Applied Biotechnology. 7-25 Mira st, Krasnogorsk, 143444, Moscovskaya oblast (mkr. Opalikha), Russian Federation. Tel: 8 (915) 054 79 73. E-mail: email@example.com
Published: June 25, 2013
The microbiological synthesis of [2H]amino acids was performed by the conversion of low molecular weight substrates ([U-2H]MeOH and 2H2O) using the Gram-positive aerobic facultative methylotrophic bacterium Brevibacterium methylicum, an L-phenylalanine producer, realizing the NAD+ dependent methanol dehydrogenase (EC 18.104.22.168) variant of the ribulose-5-monophosphate (RuMP) cycle of carbon assimilation. In this process, the adapted cells of the methylotroph with enhanced growth characteristics were used on a minimal salt medium M9, supplemented with 2% (v/v) [U-2H]MeOH and an increasing gradient of 2Н2O concentration from 0; 24.5, 49.0; 73.5 up to 98% (v/v) 2Н2O. Alanine, valine, and leucine/isoleucine were produced and accumulated exogeneously in quantities of 5–6 mmol, in addition to the main product of biosynthesis. This method enables the production of [2Н]amino acids with different degrees of deuterium enrichment, depending on the 2Н2O concentration in the growth medium, from 17 at.% 2Н (on the growth medium with 24.5 % (v/v) 2Н2О) up to 75 at.% 2Н (on the growth medium with 98 % (v/v) 2Н2О). This has been confirmed with the data from the electron impact (EI) mass spectrometry analysis of the methyl ethers of N-dimethylamino(naphthalene)-5-sulfochloride [2H]amino acids under these experimental conditions.
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