MOLECULAR-CLONING AND CHARACTERIZATION OF THE GENE ENCODING RIBOSYL)-FORMIMINO]-5-AMINOIMIDAZOLE-4-CARBOXAMIDE RIBONUCLEOTIDE (BBM-II) ISOMERASE FROM ARABIDOPSIS-THALIANA

We have isolated an Arabidopsis BBM II isomerase cDNA from an Arabidop sis cDNA library, by means of functional complementation of the E. col i hisA mutant strain HfrG6. The isolated cDNA encodes a polypeptide of 304 amino acids with a calculated molecular weight of 33 363. Sequenc e comparison with the HIS6 proteins of yeasts revealed that Arabidopsi s BBM II isomerase contains an N-terminal extension of approximately 4 0 amino acids that shows the general properties of chloroplast transit peptides. This finding is consistent with the localization of other h istidine biosynthetic enzymes, such as imidazoleglycerolphosphate dehy dratase and histidinol dehydrogenase, in the chloroplasts in higher pl ants. The primary structure of the mature protein was 50% and 42% iden tical, respectively, to the HIS6 proteins of Schizosaccharomyces pombe and Saccharomyces cerevisiae, respectively, while no prominent sequen ce similarity to the bacterial BBM II isomerase was found. That the is olated Arabidopsis cDNA actually encodes a functionally active BBM II isomerase activity was confirmed in an in vitro enzyme assay using a c rude extract prepared from strain HfrG6 transformed with the Arabidops is BBM II isomerase cDNA.