MOLECULAR CLONING OF HUMAN UMP SYNTHASE

Research Project

Project/Area Number	61480220
Research Category	Grant-in-Aid for General Scientific Research (B)
Allocation Type	Single-year Grants
Research Field	Pediatrics
Research Institution	Nagoya City University Medical School
Principal Investigator	WADA Yoshiro Nagoya City University Medical School, 医学部, 教授 (30004849)
Co-Investigator(Kenkyū-buntansha)	HIDEKO Morishita Nagayo City University Medical School, 医学部, 助手 (90174416) MAKOTO Yazaki Nagoya City University Medical School, 医学部, 助手 (80133479)
Project Period (FY)	1986 – 1987
Project Status	Completed (Fiscal Year 1987)
Budget Amount *help	¥4,200,000 (Direct Cost: ¥4,200,000) Fiscal Year 1987: ¥500,000 (Direct Cost: ¥500,000) Fiscal Year 1986: ¥3,700,000 (Direct Cost: ¥3,700,000)
Keywords	Hereditary oroticaciduria / Orotate phosphoribosyltransferase / OMP decarboxylase / オペロン / UMP合成酵素 / ODCのcDNA / オロット酸ホスホリボシールトランスフェラーゼ / オロチジル酸脱水素酵素 / cDNA
Research Abstract	In order to clarify the cause of hereditary orotic aciduria, UMP synthase (a multifunctional protein having both orotate phosporibosyltransferase and OMP decarboxylase activities) was purified from normal human erythrocytes. The enzyme protein had a molecular weight of 51,000. Then a cDNA fragment of mouse OMP decarboxylase was isolated from a <lambda>gtll mouse spleen cDNA library by the use of a synthesized oligonucleotide probe. Screening of a <lambda>gtll human placenta cDNA library yielded two positive clones which hybridized strongly to the mouse cDNA fragment. Analysis of the nucleotide sequence of the entire cDNA insert (1.7 kb) of one of the clones indicated that it contained an open reading frame (1,444 bp) encoding a protein with a molecular weight of 52,000. The deduced amino acid sequence of the 3'-half of the insert showed 89% homology with that deduced from Ehrlich ascites carcinoma OMP decarboxlase cDNA. Northern blot analysis revealed a presence of a single band of approximately 1.8 kb, which suggests that the cloned cDNA contains the whole message for human UMP synthase.