Analysis of 0.5-kilobase-pair repeats in the Mycoplasma hominis lmp gene system and identification of gene products

Abstract

Mycoplasma hominis, an opportunistic pathogenic bacterium of humans, has a small genome of 700 kb. Despite this, multiple copies of gene sequences with similarities to the structural gene (lmp1) of a 135-kDa surface-located membrane protein (Lmp1) have been identified on the genome of M. hominis PG21 (lmp2, lmp3, and lmp4). The distance between the lmp1-lmp2 region and the lmp3-lmp4 region was more than 110 kb. lmp3-lmp4 of M. hominis PG21 was sequenced and found to contain two putative genes. The gene region of 6.5 kb contained a 5 unique region and a 3 unique region separated by 9 0.5-kb repeats with 51 to 90% similarity to 10 similar repeats found in the lmp1-lmp2 region. The 0.5-kb DNA repeats thus comprised about 1% of the entire genome. In both regions, a base change in one of the repeats gave rise to a stop codon, and thereby lmp2 and lmp4 occurred. By PCR amplification of reverse-transcriptase-generated cDNA it was shown that all four genes were transcribed. By use of Lmp-specific antibodies we showed that both lmp1 and lmp3 were translated into proteins (Lmp1 and Lmp3). Each of the four lmp genes represented by their unique cloned segments was used as a probe to analyze the presence, distribution, and organization of the genes within the genome in 13 M. hominis isolates. The repetitive element was detected at one or two locations on the chromosome for all isolates. The lmp3-specific element was present in all isolates, and lmp1-and lmp2-specific elements were present in all but one isolate. The lmp4-specific element was present in about half the isolates tested. For five M. hominis isolates the chromosomal location of the lmp genes was mapped.Mycoplasmas are the smallest prokaryotes capable of selfreplication. They have small genomes of 600 to 1,600 kb with limited biosynthetic capabilities (2, 21). To meet their nutritional requirements, these bacteria rely upon a parasitic lifestyle on mucosal surfaces in humans. Mycoplasma hominis is an opportunistic pathogen causing gynecological infections. Extragenital infections with M. hominis are recognized with increasing frequency (3).M. hominis PG21 possesses a 135-kDa surface-located membrane protein Lmp1 identified by monoclonal antibody (MAb) 552 and encoded by the lmp1 gene (14). A membrane protein recognized by MAb 552 is generally present in M. hominis. Of 26 M. hominis isolates tested, MAb 552 reacted with a single band of 93 kDa in 22 isolates. In the remaining four isolates including M. hominis PG21, MAb 552 reacted with a ladder of polypeptides ranging in size from 93 to 153 kDa (5).The gene coding for Lmp1 is located 253 bp from the tuf gene (15). It covers 4,032 bp, equivalent to 1,344 amino acids and a molecular size of 147 kDa. lmp2 is a gene situated immediately downstream from lmp1 covering 2,886 bp. Following the first 726 bp from the start codon of lmp1, nine and a half similar direct repeats of 471 nucleotides are situated. The stop codon of lmp1 is located within repeat 8. Following the stop codon is a methionine codon that initiates the ...

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