Isolation and characterization of a histidine biosynthetic gene in Arabidopsis encoding a polypeptide with two separate domains for phosphoribosyl-ATP pyrophosphohydrolase and phosphoribosyl-AMP cyclohydrolase.
Phosphoribosyl-ATP pyrophosphohydrolase (PRA-PH) and phosphoribosyl-AMP cyclohydrolase (PRA-CH) are encoded by HIS4 in yeast and by hisIE in bacteria and catalyze the second and the third step, respectively, in the histidine biosynthetic pathway. By complementing a hisI mutation of Escherichia coli with an Arabidopsis cDNA library, we isolated an Arabidopsis cDNA (At-IE) that possesses these two enzyme activities. The At-IE cDNA encodes a bifunctional protein of 281 amino acids with a calculated molecular mass of 31,666 D. Genomic DNA-blot analysis with the At-IE cDNA as a probe revealed a single-copy gene in Arabidopsis, and RNA-blot analysis showed that the At-IE gene was expressed ubiquitously throughout development. Sequence comparison suggested that the At-IE protein has an N-terminal extension of about 50 amino acids with the properties of a chloroplast transit peptide. We demonstrated through heterologous expression studies in E. coli that the functional domains for the PRA-CH (hisI) and PRA-PH (hisE) resided in the N-terminal and the C-terminal halves, respectively, of the At-IE protein.
Fujimori
K
K
Takarazuka Research Institute, Novartis Pharma K.K., 10-66 Miyuki-cho, Takarazuka 665-8666, Japan.
Ohta
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D
D000595:Amino Acid Sequence; D000619:Aminohydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D001483:Base Sequence; D020134:Catalytic Domain / Q000235:genetics; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D018744:DNA, Plant / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D017343:Genes, Plant; D005816:Genetic Complementation Test; D006639:Histidine / Q000096:biosynthesis* / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D011755:Pyrophosphatases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D018744:DNA, Plant; D010940:Plant Proteins; D006639:Histidine; D000619:Aminohydrolases; C022383:phosphoribosyl-AMP cyclohydrolase; D011755:Pyrophosphatases; C064815:phosphoribosyl-ATP pyrophosphatase