The amino acid L-serine is required for pharmaceutical purposes, and the availability of a sugar-based microbial process for its production is desirable. However, a number of intracellular utilization routes prevent overproduction of L-serine, with the essential serine hydroxymethyltransferase (SHMT) (glyA) probably occupying a key position. We found that constructs of Corynebacterium glutamicum strains where chromosomal glyA expression is dependent on P tac and lacI Q are unstable, acquiring mutations in lacI Q , for instance. To overcome the inconvenient glyA expression control, we instead considered controlling SHMT activity by the availability of 5,6,7,8-tetrahydrofolate (THF). The pabAB and pabC genes of THF synthesis were identified and deleted in C. glutamicum, and the resulting strains were shown to require folate or 4-aminobenzoate for growth. Whereas the C. glutamicum ⌬sdaA strain (pserACB) accumulates only traces of L-serine, with the C. glutamicum ⌬pabABC⌬sdaA strain (pserACB), L-serine accumulation and growth responded in a dose-dependent manner to an external folate supply. At 0.1 mM folate, 81 mM L-serine accumulated. In a 20-liter controlled fed-batch culture, a 345 mM L-serine accumulation was achieved. Thus, an efficient and highly competitive process for microbial L-serine production is available.L-Serine is a nonessential amino acid but plays an important role in stabilizing the blood sugar concentration in the liver (16). It relates, furthermore, to many other substances, including sphingosine and the phosphatides, which are part of the myelin covering of the nerves, as well as the formation of activated C 1 units used for a number of anabolic processes (20). Therefore, L-serine is present in selected infusion solutions and also has other applications. For instance, it is an ingredient of skin lotions to ensure a proper hydration status. The total annual demand for L-serine is estimated to be 300 tons (5).The production processes currently used still rely on the extraction of L-serine from protein hydrolysates or from molasses, as well as on the enzymological conversion of glycine plus a C 1 compound, like methanol, to L-serine. The latter uses the reverse reaction of the serine hydroxymethyltransferase (SHMT) (6). Thus, an enzymatic system has been designed to convert glycine plus formaldehyde to L-serine (4). The cellular systems assayed employed, among other things, resting cells of methanol-utilizing bacteria, such as Hyphomicrobium methylovorum, where L-serine formation from glycine plus methanol was achieved (6). In such a system, up to 45 g liter Ϫ1 L-serine accumulation was possible, but only at a glycine yield of 50%, thus making the system less attractive. Also, alginate-entrapped cells of Corynebacterium glycinophilum were used for L-serine formation from glycine (21). It is self-evident that it would be most profitable to directly convert cheap sugar into L-serine. Although microbial processes for amino acid production are in general advancing quickly, attempts to develop L-serin...
