IDENTIFICATION OF A PUTATIVE INFC-RPMI-RPLT OPERON FLANKED BY LONG INVERTED REPEATS IN MYCOPLASMA-FERMENTANS (INCOGNITUS STRAIN)

A specific 1542-bp DNA fragment was amplified from Mycoplasma fermenta ns (incognitus strain) using a unique 23-nucleotide (nt) synthetic deo xyribonucleotide (oligo) (5'-TCCAAAAAGTCCGGAATTTGGGG) as the primer pa ir in the polymerase chain reaction (PCR). The 23-nt sequence is part of the 29-bp terminal inverted repeat (IR) which forms the left potent ial stem-and-loop (s&1) structure of the previously identified M. ferm entans insertion-sequence(IS)-like genetic element [Hu et al., Gene 93 (1990) 67-72]. The amplified DNA was cloned and sequenced. A pair of 27-bp IR containing the 23-nt synthetic oligo was identified at both t ermini. Between the IR, there are four potential open reading frames ( ORFs) which are arranged adjacent to each other in the order, ORF-1, O RF-2, ORF-3 and ORF-4, with parts of ORF-1 and ORF-2 overlapping. The deduced amino acid (aa) sequences of ORF-2, ORF-3 and ORF-4 are 34 to 60% identical to the translation initiation factor IF3 (encoded by the infC gene), ribosomal proteins L35 (rpmI gene) and L20 (rplT gene) of Escherichia coli and Bacillus stearothermophilus, respectively. In ba cteria, the infC-rpmI-rplT genes are organized to function as an opero n. There are multiple sites with promoter-like sequences identified up stream from the putative infC gene in the mycoplasma closely resemblin g the gene arrangement in the bacterial operon. All three genes of ORF -2, ORF-3 and ORF-4 are preceded individually by a strong appropriatel y spaced (7 and 10 bp) putative Shine-Dalgamo sequence (5'-AAGGA). In addition, ORF-2 uses the unusual triplet, ATT, as the start codon, the same as that for infC in the bacterial operon. Thus, the cluster of g enes (ORF-2, ORF-3 and ORF-4) is identified as a putative mycoplasma i nfC-rpmI-rplT operon. Most interestingly, our study reveals that this operon potentially constitutes a part of a mobile genetic element in t he incognitus strain of M. fermentans.