Disruption of the SHM2 gene, encoding one of two serine hydroxymethyltransferase isoenzymes, reduces the flux from glycine to serine in Ashbya gossypii.
Riboflavin overproduction in the ascomycete Ashbya gossypii is limited by glycine, a precursor of purine biosynthesis, and therefore an indicator of glycine metabolism. Disruption of the SHM 2 gene, encoding a serine hydroxymethyltransferase, resulted in a significant increase in riboflavin productivity. Determination of the enzyme's specific activity revealed a reduction from 3 m-units/mg of protein to 0.5 m-unit/mg protein. The remaining activity was due to an isoenzyme encoded by SHM 1, which is probably mitochondrial. A hypothesis proposed to account for the enhanced riboflavin overproduction of SHM 2-disrupted mutants was that the flux from glycine to serine was reduced, thus leading to an elevated supply with the riboflavin precursor glycine. Evidence for the correctness of that hypothesis was obtained from (13)C-labelling experiments. When 500 microM 99% [1-(13)C]threonine was fed, more than 50% of the label was detected in C-1 of glycine resulting from threonine aldolase activity. More than 30% labelling determined in C-1 of serine can be explained by serine synthesis via serine hydroxymethyltransferase. Knockout of SHM 1 had no detectable effect on serine labelling, but disruption of SHM 2 led to a decrease in serine (2-5%) and an increase in glycine (59-67%) labelling, indicating a changed carbon flux.
D000595:Amino Acid Sequence; D002250:Carbon Radioisotopes / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D018390:Gene Targeting; D005800:Genes, Fungal; D005998:Glycine / Q000378:metabolism*; D012696:Glycine Hydroxymethyltransferase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007527:Isoenzymes / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012256:Riboflavin / Q000096:biosynthesis; D004718:Saccharomycetales / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012694:Serine / Q000378:metabolism*; D013912:Threonine / Q000378:metabolism
D002250:Carbon Radioisotopes; D005656:Fungal Proteins; D007527:Isoenzymes; D013912:Threonine; D012694:Serine; D012696:Glycine Hydroxymethyltransferase; D005998:Glycine; D012256:Riboflavin