Isolation and partial characterization of a 5'-nucleotidase specific for orotidine-5'-monophosphate.

Abstract

A previously unknown 5'nucleotidase (5'-ribonucleotide phosphohydrolase, EC 3.1.3.5) (5'-Nase) specific for orotidine 5'-monophosphate (OMP) hs been discovered. This enzyme orotidine 5'-monophosphate phosphohydrolase (OMPase), was isolated from mouse liver microsomes as a separate entity from the nonspecific 5'-Nase. OMPase was partially purified and is shown to cleave OMP to orotidine and inorganic phosphate. The enzyme has negligible activity towards UMP, CMP, dTMP, AMP, IMP, GMP, XMP, 6-azauridine 5'-monophosphate, 1-beta-D-ribofuranosylbarbituric acid 5'-monophosphate (BMF), 2'-UMP, 3'-UMP, 2'-AMP, 3'-AMP, ribose 5-phosphate and beta-glycerophosphate, all of which--with the exception of the 2' or 3' monophosphates, ribose 5'-phosphate, and beta-glycerophosphate--are substrates for 5'-Nase. Both enzymes are inhibited by NaF, but only OMPase is inhibited by SF reagents. OMPase is not inhibited by orotidine, orotate, BMP, concanavalin A, or tetramisole (an alkaline phosphatase inhibitor). OMPase had a Mr 53,000, Km value of 1 mM for OMP, and Vmax value of 49 nmol/min . mg of protein at the present stage of purification. OMPase activity has also been detected in various mammalian tissues including normal human tissues, human tumor xenografts, lymphocytes, and rat liver. OMPase may be responsible, in part, for the low levels of intracellular "free" OMP and for orotidine accumulation in cells treated with 6-azauridine and patients suffering from aortic aciduria.

• Authors: M H El Kouni, S Cha
• Journal: Proceedings of the National Academy of Sciences of the United States of America (Proc Natl Acad Sci U S A) Vol. 79 Issue 4 Pg. 1037-41 (Feb 1982) ISSN: 0027-8424 [Print] United States
• PMID: 6280163 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Uracil Nucleotides
• Concanavalin A
• orotidylic acid
• Sodium Fluoride
• Dithionitrobenzoic Acid
• Uridine Monophosphate
• Pentosyltransferases
• Orotate Phosphoribosyltransferase
• Nucleotidases
• 5'-Nucleotidase
• Magnesium

Topics

• 5'-Nucleotidase
• Animals
• Concanavalin A (pharmacology)
• Dithionitrobenzoic Acid (pharmacology)
• Kinetics
• Magnesium (pharmacology)
• Male
• Mice
• Microsomes (enzymology)
• Nucleotidases (isolation & purification, metabolism)
• Orotate Phosphoribosyltransferase (isolation & purification)
• Pentosyltransferases (isolation & purification)
• Rats
• Sodium Fluoride (pharmacology)
• Substrate Specificity
• Tissue Distribution
• Uracil Nucleotides (metabolism)
• Uridine Monophosphate (analogs & derivatives, metabolism)