MOLECULAR-CLONING AND NUCLEOTIDE-SEQUENCES OF THE GENES FOR 2 ESSENTIAL PROTEINS CONSTITUTING A NOVEL ENZYME-SYSTEM FOR HEPTAPRENYL DIPHOSPHATE SYNTHESIS

The genes encoding two dissociable components essential for Bacillus s tearothermophilus heptaprenyl diphosphate synthase ans-hexaprenyl-diph osphate:isopentenyl-diphosphate hexaprenyl-trans-transferase, EC 2.5.1 .30) were cloned, and their nucleotide sequences were determined. Sequ ence analyses revealed the presence of three open reading frames withi n 2,350 base pairs, designated as ORF-1, ORF-2, and ORF-3 in order of nucleotide sequence, which encode proteins of 220, 234, and 323 amino acids, respectively. Deletion experiments have shown that expression o f the enzymatic activity requires the presence of ORF-1 and ORF-3, but ORF-2 is not essential. As a result, this enzyme was proved genetical ly to consist of two different protein components with molecular masse s of 25 kDa (Component I) and 36 kDa (Component II), encoded by two of the three tandem genes. The protein encoded by ORF-1 has no similarit y to any protein so far registered. However, the protein encoded by OR F-3 shows a 32% similarity to the farnesyl diphosphate synthase of the same bacterium and has seven highly conserved regions that have been shown typical in prenyltransferases (Koyama, T., Obata, S., Osabe, M., Takeshita, A., Yokoyama, K., Uchida, M., Nishino, T., and Ogura, K. ( 1993) J. Biochem. (Tokyo) 113, 355-363).