Production of glutamate and stereospecific flavors, (S)-linalool and (+)-valencene, by Synechocystis sp. PCC6803
Section snippets
Strains and plasmids
Details of the strains and plasmids used in this study are summarized in Table 1 (15,16). Our laboratory strain GT-A, which was derived from the glucose-tolerant (GT) derivative of Synechocystis sp. PCC 6803 (17), was used as the parental strain of all the mutants constructed in this study. Escherichia coli JM109 was used for plasmid construction and propagation.
Plasmid construction
The sequences of the primers used in this study are shown in Table S1. To construct a plasmid expressing E. coli YbjL (18), the ybjL
l-Glutamate production
As a preliminary test, l-glutamate (0.25–1.0 g/L) was added to the culture medium of Synechocystis sp. PCC6803. Addition of l-glutamate increased the aggregation of the cells, however, consumption of l-glutamate was not observed. Thus, l-glutamate uptake activity in Synechocystis sp. PCC6803 was estimated to be extremely low. We also assumed that the activity to export l-glutamate is low in Synechocystis sp. PCC6803 (Fig. 1A), as is the case of sucrose and lactic acid (5). We hence introduced
Discussion
l-Glutamate is negatively charged at neutral pH, thus export across the membrane typically requires an efflux carrier. In l-glutamate producing bacterium, Corynebacteirum glutamicum, mechanosensitive channels, MscCG (originally named NCgl1221), function as a major glutamate exporter (36). In enteric bacteria like E. coli, we have found that YbjL functions as an exporter of acidic substances having carboxyl groups, such as succinate and l-glutamate (18). The introduction of a single gene
Acknowledgments
We thank Prof. Masahiko Ikeuchi and the late Dr. Takafumi Midorikawa of the Department of Life Sciences, University of Tokyo, for kindly providing Synechocystis sp. PCC6803 GT strains and plasmids, and Dr. Shigeo Suzuki, Mr. Yuta Nagasaka, Mr. Wataru Hoshino, Dr. Shuhei Hashiro, and Mr. Hiroaki Fukada of Ajinomoto Co., Inc. for technical advice and support.
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