Orotate phosphoribosyltransferase (EC 2.4.2.10) and
orotidine 5'-monophosphate decarboxylase (EC 4.1.1.23) are the final two of six enzymatic steps required in the de novo biosynthesis of
uridine 5'-monophosphate (
UMP). Earlier work of this laboratory showed that, in the mouse Ehrlich
ascites carcinoma, both of these enzymatic activities were contained on the single multifunctional
polypeptide chain,
UMP synthase. We report here that the placenta provided an available human source for
UMP synthase with 40-fold higher
orotate phosphoribosyltransferase and
orotidine 5'-monophosphate decarboxylase specific activities than erythrocytes, a human source previously used by others. By using the placenta as a source of
UMP synthase and by developing a novel purification procedure different from that used in the purification of
UMP synthase from the Ehrlich
ascites carcinoma (the only other homogeneous preparation of a mammalian
UMP synthase), we achieved the purification of human
UMP synthase to apparent homogeneity. This represents the first publication to homogeneity of
UMP synthase from a human source as well as from a source other than malignant cell lines. Contrary to earlier reports human placental
UMP synthase was found to be a multifunctional
protein containing both enzymatic activities on a single
polypeptide of 51,000 molecular weight. Preliminary characterization of the human placental
UMP synthase showed it to be similar to
UMP synthase from the Ehrlich
ascites carcinoma in subunit molecular weight, native molecular weight,
isozyme pattern (although not absolute pI values), pH optima of enzymatic activities, and kinetic constants for
orotidine 5'-monophosphate (Km) and
6-azauridine 5'-monophosphate (Ki) at the
decarboxylase site.