Tyrosine Decarboxylase: II. Pyridoxine-deficient Medium for Apoenzyme Production.

Previous communications from this laboratory have established the function of pyridoxine derivatives' in the tyrosine decarboxylase system of Streptococcus faecalis, and a synthetic medium has been described for the production of cells active in this function (Bellamy and Gunsalus, 1943, 1944). Cells grown in media deficient in,pyridoxine derivatives have been shown to contain the tyrosine decarboxylating enzyme but not the coenzyme (Gunsalus and Bellamy, 1944a). Pyridoxal (Snell, 1944) will activate this enzyme in the living cells (Gunsalus and Bellamy, 1944b) but dried cell preparations from them are active only if adenosine triphosphate (ATP) is supplied in addition to pyridoxal (Gunsalus, Bellamy, and Umbreit, 1944). The growth of cells deficient in coenzyme appeared to offer a simpler method for the preparation of the decarboxylase apoenzyme than would the more conventional method of isolation and resolution of the enzyme. Therefore, the present work was undertaken. The main objective has been the production of the greatest possible yield of tyrosine decarboxylase apoenzyme with minimum coenzyme content per unit of medium. This differs from the previous study in which the primary purfiose was to find which vitamin or vitamins function in the tyrosine system and to produce as active a tyrosine decarboxylase as possible (Bellamy and Gunsalus, 1944). The method of apoenzyme production utilized the discovery of Snell and Guirard (1943) that an excess of alanine will support growth of Streptococcus faecalis, strain R, in the absence of pyridoxine derivatives. The lack of coenzyme in cells grown with alanine indicates that vitamin B6 analogues convertible into coenzyme are not synthesized from alanine during growth. This constitutes an exception to the working hypothesis used by Snell and Guirard (1943), who suggest that alanine might be used as an essential nucleus for the synthesis of pyridoxine. However, evidence of synthesis of pyridoxine derivatives by other lactic acid bacteria has been reported by Bohonos, Hutchings, and Peterson (1942). In the present paper, a medium for the production of a very active tyrosine decarboxylase apoenzyme in Streptococcusfaecalis, strain R, is reported, and some of the factors affecting its production are discussed. These cells respond quantitatively to pyridoxal, and dried cell preparations from them respond to pyridoxal in the presence of ATP.