Isolation and sequencing of a genomic DNA clone containing the 3′ terminus of the 6-methylsalicylic acid polyketide synthetase gene of Penicillium urticae

Abstract

A 7.7-kilobase (kb) Penicillium urticae genomic DNA fragment containing the 3' terminus of the 6-methylsalicylic acid polyketide synthetase gene was cloned using a 41-mer mixed oligodeoxynucleotide probe which was based on a cyanogen bromide cleavage peptide of 35 amino acids obtained from pure synthetase. Nucleotide sequence analysis of a 2.2-kb region of the cloned fragment revealed a large open reading frame of 1866 bases which was devoid of introns and which corresponded to amino acids of the carboxyl terminus of the enzyme. This was followed by a putative transcription termination--polyadenylation signal. A putative acyl carrier protein domain at the 3' terminus was preceded by a beta-ketoreductase domain. These functionalities were identified by amino acid sequences known to be characteristic of the active sites of fatty acid synthetase functional domains. Their relative positions contrast with those in yeast and P. urticae fatty acid synthetase genes where the two functional domains are located at the 5' terminus and in reverse order. Furthermore, amino acid sequence identities indicated a striking homology with vertebrate rather than either yeast or P. urticae fatty acid synthetases.