Patente De Glutamato

JOURNAL OF BIOSCIENCE AND BIOENGINEERING Vol. 98, No. 3, 211–213. 2004

Enhanced Glutamic Acid Production of Brevibacterium sp. with Temperature Shift-Up Cultivation
SUN-UK CHOI,1§ TAKUYA NIHIRA,1 AND TOSHIOMI YOSHIDA1§§*
International Center for Biotechnology, Osaka University, 2-1 Yamadaoka, Suita, Osaka 565-0871, Japan1
Received 18 May 2004/Accepted 27 May 2004

For cells ofBrevibacterium sp. under conditions of biotin limitation, the efflux of metabolites through the cell membrane was affected by temperature. After the temperature shift-up from 30°C to 38°C, both the specific production rate of glutamic acid and the excretion of intracellular materials, such as glucose-6-phosphate and nucleotides, were increased simultaneously. For the production of glutamic acid, not only theyield but also the specific production rate was increased by the temperature shift-up.
[Key words: glutamic acid, efflux, biotin limitation, shift-up]

Excretion of glutamic acid by cells of Brevibacterium sp. and Corynebacterium sp. has been investigated to improve the productivity of glutamic acid fermentation, based on the finding that the efflux of glutamic acid, and thus the yield ofglutamic acid, can be enhanced by various treatments which seem to alter the bacterial membrane. These treatments include biotin limitation (1), treatment with detergents (2), addition of penicillin (3), and fermentation using glycerol or oleic acid auxotrophs (4, 5), with biotin limitation having been used effectively in the industry-scale production of glutamic acid. With respect to the excretionmechanism of glutamic acid, three hypotheses which are still under debate have been proposed, namely, (i) a leak model (1, 6), (ii) an inversion model (7), and (iii) a model employing a specific transport system for glutamic acid, based on a general efflux model in bacteria (8). The first hypothesis (the leak model) has been widely accepted to explain the excretion of glutamic acid under biotinlimitation. This model is based on the observation that biotin deficiency suppresses fatty acid synthesis, which alters the structures/fluidity of the plasma membrane by decreasing the lipid content together with the composition change of both phospholipids and fatty acids (1, 6, 9). Previously, we confirmed that cultivation temperature affects the production of glutamic acid, and estimated thatcultivation temperature may affect the efflux of glutamic acid (10). Analysis of the lipid composition of the cell membrane (11, 12) indicated that the degree of fluidity depends heavily on lipid composition and temperature. When the cultivation was performed at low temperature, cells adapted to the temperature by increasing the amount of unsaturated
* Corresponding author. e-mail:tyoshida0801@icb.osaka-u.ac.jp phone: +66-2-661-6453 fax: +66-2-661-6454 Present address: § Division of Food and Biotechnology, Kyungnam University, Masan 631-701, Korea and §§ JSPS Bangkok Office, 113 TWY Office Center, 10th Fl., Serm-mit Tower, 159 Sukhumvit Soi 21, Bangkok 10110, Thailand.
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fatty acid moieties containing cis-double bonds in the cell membrane to increase the fluidity of the lipid bilayerthrough which the efflux of small materials and nutrients increased to the level similar to that under the normal cultivation temperature. Although the cells have the ability to normalize membrane fluidity under different temperatures through gradual adaptation (11), the membrane fluidity under sudden temperature change, and hence, the efflux through the cell membrane, could be determined by thetemperature if the cell membrane is defective and structurally weak due to biotin limitation (1, 6, 9, 13). In this study, we showed that the efflux of glutamic acid from cells of Brevibacterium sp. is affected by temperature. The yield, as well as the specific production rate, of glutamic acid was also increased by temperature shift-up. Brevibacterium sp. strain TC452 from our stock was used...