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The study of the subcellular distribution of the enzymes involved in folate-coenzyme-mediated transfer of one-carbon groups in Saccharomyces cerevisiae suggests the existence of two parallel enzyme systems, one mitochondrial and the other cytoplasmic. All the enzymes assayed in the present study, serine transhydroxymethylase, dihydrofolate reductase, 5,lO-methylenetetrahydrofolate dehydrogenase, 10-formyltetrahydrofolate synthetase, 5,lO-methylenetetrahydrofolate reductase and thymidylate synthetase were found to be in both cytoplasm and mitochondria. Two of the mitochondrial enzymes, serine transhydroxymethylase and dihydrofolate reductase, were found to be different by several criteria from their cytoplasmic counterparts.By the use of three nuclear gene mutations ade3, tmp3 and serl, and different combinations of these three mutations, the physiological role of the cytoplasmic enzymes for the biosynthesis of adenine, histidine, methionine, thymidylate and serine was studied. The response of auxotrophes to an external source of tetrahydrofolate, i. e. 5-formyl tetrahydrofolic acid, was also examined. The mitochondrial set of enzymes might be responsible for the de novo formation of tetrahydrofolate, since the loss of the three mitochondrial enzyme activities, serine transhydroxymethylase, thymidylate synthetase and dihydrofolate reductase, due to the mutation tmp3, leads to a generalized requirement for thymidylate, methionine, adenine and histidine. The ade3 mutation in which the cytoplasmic activities of the 10-formyltetrahydrofolate synthetase, 5,lO-methenyltetrahydrofolate cyclohydrolase and 5,lO-methylenetetrahydrofolate dehydrogenase are lost, confers only adenine and histidine auxotrophy, showing that the ade3 proteins are synthesized in, and only function in, the cytoplasm. The mitochondrial counterparts of these enzymes may function solely in the formation of the 10-formyl tetrahydrofolate needed for the transformylation of Met-tRNA involved in mitochondrial protein synthesis.A study of a nuclear mutation in Saccharomyces cerevisiae, tmp3, leading concomittantly to multiple nutritional requirements for thymidylate, methionine, adenine and histidine and to the petite phenotype, was found to have a decreased activity of serine transhydroxymethylase, but a wild-type level of two other tetrahydrofolate-linked enzymes, 10-formylte- trahydrofolate-linked enzymes, 10-formyltetrahydrofolate synthetase and 5,1O-methylenetetrahydrofolate dehydrogenase [l]. Further study showed that wildtype strains of yeast have two forms of serine transhydroxymethylase, one cytoplasmic and the other mitochondrial, separable by TEAE-cellulose chromatography. The mutant tmp3 has lost the mitochondrial form [2].Serine transhydroxymethylase may be the major physiological source of 5,lO-methylene tetrahydrofolate needed for the formation of methionine, thymidylate and adenine. Loss of the mitochondrial enzyme, in tmp3, which leads to that multiple requirement, implies that either the mitochondrial serine transhydroxymethylase is the...

Two forms of serine transhydroxymethylase, separable by TEAE‐cellulose chromatography, are present in wild‐type extracts of Saccharomyces cerevisiae. One of these forms is mitochondrial and the other one cytoplasmic. The mitochondrial form is absent in the mutant tmp3.

A chromosomal linkage map of ten markers was established for Streptomyces ambofaciens by the four-factor cross method and allele-gradient analysis. Mutants were obtained by nitrous acid treatment as well as UV mutagenesis. The fertility of crosses was enhanced over 100-fold by pSAM2, a plasmid present in some strains of S. ambofaciens, and over 1000-fold by the conjugative plasmid pIJ303.