Purification and characterization of Arabidopsis ornithine transcarbamoylase (OTCase), a member of a distinct and evolutionarily-conserved group of plant OTCases

详细信息    查看全文

• 作者：Slocum ; Robert D. ; Nichols III ; Harry F. ; Williamson ; Cynthia L.
• 关键词：Arabidopsis thaliana ; argF ; arginine biosynthesis ; carbamoyl phosphate ; ornithine transcarbamoylase ; ATCase ; aspartate transcarbamoylase ; carbamoyl-P ; carbamoyl phosphate ; OTCase ; ornithine transcarbamoylase ; PALO ; N-(phosphonacetyl)-L-ornithine ; pOTCase
• 刊名：Plant Physiology and Biochemistry
• 出版年：2000
• 出版时间：April, 2000
• 年：2000
• 卷：38
• 期：4
• 页码：279-288
• 全文大小：633 K

文摘

We affinity-purified an ornithine transcarbamoylase (carbamoyl phosphate:L-ornithine carbamoyltransferase; EC 2.1.3.3) 676-fold to near homogeneity from leaf tissues of Arabidopsis thaliana L. cv. Columbia. The purified OTCase protein exhibited a molecular mass of 37 kDa on SDS-PAGE gels and exhibited a pI = 6.8. A 41-kDa polypeptide was immunoprecipitated from Arabidopsis leaf poly(A)⁺ RNA in vitro translation products by pea OTCase antiserum. This precursor OTCase (pOTCase) is the predicted size (41 170 Da) for a polypeptide encoded by an Arabidopsis OTCase cDNA. Characteristics of N-terminal residues of the deduced amino acid sequence of this pOTCase suggest that it is a chloroplast-targeted protein. The sequences of plant OTCases suggest that they represent a distinct and evolutionarily-conserved group of OTCases. No evidence was found for OTCase isoenzymes in Arabidopsis leaf tissues. The Arabidopsis pOTCase was poorly-expressed in Escherichia coli strain TB-2, an OTCase-deficient mutant, and did not complement the mutant on arginine-minus selection medium.