A Brassica cDNA clone encoding a bifunctional hydroxymethylpyrimidine kinase/thiamin-phosphate pyrophosphorylase involved in thiamin biosynthesis

被引：27

作者：

Kim Y.S. ^{[1
]}

Nosaka K. ^{[2
]}

Downs D.M. ^{[3
]}

Kwak J.M. ^{[1
]}

Park D. ^{[1
]}

Chung II K. ^{[4
]}

Nam H.G. ^{[1
]}

机构：

[1] Department of Life Science, School of Environmental Engineering, Pohang Univ. of Sci. and Technology, Pohang, Kyungbuk

[2] Department of Biochemistry, Kyoto Pref. University of Medicine, Kamigyo-ku

[3] Department of Bacteriology, University of Wisconsin-Madison, Madison

[4] Department of Horticultural Science, Catholic University of Taegu-Hyosung, Kyungsan, Kyungbuk

来源：

Plant Molecular Biology | 1998年 / 37卷 / 6期

基金：

美国国家卫生研究院;

关键词：

Bifunctional enzyme; Brassica napus; cDNA; Hydroxymethylpyrimidine phosphate kinase; Thiamin; Thiamin phosphate pyrophosphorylase;

D O I：

10.1023/A:1006030617502

中图分类号：

学科分类号：

摘要：

We report the characterization of a Brassica napus cDNA clone (pBTH1) encoding a protein (BTH1) with two enzymatic activities in the thiamin biosynthetic pathway, thiamin-phosphate pyrophosphorylase (TMP-PPase) and 2-methyl-4-amino-5-hydroxymethylpyrimidine-monophosphate kinase (HMP-P kinase). The cDNA clone was isolated by a novel functional complementation strategy employing an Escherichia coli mutant deficient in the TMP-PPase activity. A biochemical assay showed the clone to confer recovery of TMP-PPase activity in the E. coli mutant strain. The cDNA clone is 1746 bp long and contains an open reading frame encoding a peptide of 524 amino acids. The C-terminal part of BTH1 showed 53% and 59% sequence similarity to the N-terminal TMP-PPase region of the bifunctional yeast proteins Saccharomyces THI6 and Schizosaccharomyces pombe THI4, respectively. The N-terminal part of BTH1 showed 58% sequence similarity to HMP-P kinase of Salmonella typhimurium. The cDNA clone functionally complemented the S. typhimurium and E. coli thiD mutants deficient in the HMP-P kinase activity. These results show that the clone encodes a bifunctional protein with TMP-PPase at the C-terminus and HMP-P kinase at the N-terminus. This is in contrast to the yeast bifunctional proteins that encode TMP-PPase at the N-terminus and 4-methyl-5-(2-hydroxyethyl)thiazole kinase at the C-terminus. Expression of the BTHI gene is negatively regulated by thiamin, as in the cases for the thiamin biosynthetic genes of microorganisms. This is the first report of a plant thiamin biosynthetic gene on which a specific biochemical activity is assigned. The Brassica BTH1 gene may correspond to the Arabidopsis TH-1 gene.

引用

页码：955 / 966

页数：11

共 46 条

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