Analysis of three different repeated DNA elements present in the P1 operon of<i>Mycoplasma pneumoniae</i>: size, number and distribution on the genome

Ruland, Klaus; Wenzel, Rainer; Herrmann, Richard

doi:10.1093/nar/18.21.6311

Cited by 69 publications

(86 citation statements)

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“…The two M. pneumoniae strains tested, the clinical isolate M129 and the laboratory high-passage strain FH, show size polymorphism of DNA restriction fragments (11,47,54). In this work, we could show a size polymorphism of P65 between M129 and FH, obviously caused by an intragenetic duplication of a 54-bp sequence in orfp65 FH .…”

Section: Discussionmentioning

confidence: 54%

“…Differences in DNA and protein profiles were detected among clinical isolates of M. pneumoniae (47,54,55). On the basis of restriction fragment length polymorphism of the P1 gene M. pneumoniae isolates were classified in two distinct categories: group I, containing M129, and group II, containing the FH strain (11,54).…”

Section: Detection Of P65 In Triton X-100-insoluble Subfractionsmentioning

confidence: 99%

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The proline-rich P65 protein of Mycoplasma pneumoniae is a component of the Triton X-100-insoluble fraction and exhibits size polymorphism in the strains M129 and FH

et al. 1995

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Previously, we described the identification of a novel Mycoplasma pneumoniae M129 protein, named P65 because of its apparent molecular mass of 65 kDa estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (T. Proft and R. Herrmann, Mol. Microbiol. 13:337-348, 1994). DNA sequence analysis of the P65 open reading frame (orfp65), however, revealed an ORF encoding a protein with a molecular weight of 47,034. This discrepancy can be explained by the unusual amino acid composition of this protein. According to the deduced amino acid sequence, the N-terminal half of P65 contains several penta-and hexapeptides (DPNAY and DPNQAY) forming a proline-rich acidic domain. Secondary-structure predictions indicated ␤-sheets and turns within that region, suggesting an extended and rigid conformation. Near the C terminus of P65 the tripeptide Arg-Gly-Asp (RGD) was found. This motif is known to play an important role in binding of extracellular matrix proteins to integrins. P65 could be located exclusively to the Triton X-100-insoluble cell fraction. The results of immunofluorescence microscopy and of immunoadsorption experiments indicated that P65 carries surface-exposed regions. Mild treatment of whole cells with proteases resulted in cleavage of a limited amount of P65 molecules, suggesting either that only a small percentage of P65 molecules are exposed on the surface or that protease cleavage is hampered by a compact protein conformation or by binding of an unknown component to P65. P65 exhibits size polymorphism in M. pneumoniae M129 and FH. This is caused by an intragenetic duplication of a 54-bp sequence within the FH orfp65. As a consequence, the number of DPNAY pentapeptides increased from 9 to 12 repeats in the FH strain.Mycoplasma pneumoniae is an extracellular pathogen of the human respiratory tract (26, 46) causing histopathological changes of lung epithelial cells, usually in older children and young adults (18). A critical step in bacterial colonization of the host cells is the specific adhesion to host cell receptors, mediated by bacterial adhesins. In M. pneumoniae, the P1 adhesin (3,17,25,26) and the adhesin-related 30-kDa protein (4, 10) have been identified. Both proteins are located mainly in a tip structure that functions as the attachment organelle of the bacterium. It could be demonstrated, by nearestneighbor analysis with a hydrophilic chemical cross-linker, that the product of open reading frame 6 (ORF6) of the P1 operon (28), a 40-kDa protein and 90-kDa protein (9, 34, 50) are located in close proximity to the P1 adhesin on the cell surface (35).Scanning and transmission electron microscope analyses of M. pneumoniae cells grown on grids and pretreated with Triton X-100 revealed a rodlike tip structure and a network of filamentous strands (22,41). Krause and coworkers identified a set of high-molecular-weight proteins (HMW1 to HMW5) which play an important role in cytadherence (30,31,(51)(52)(53) and appear to be involved in formation of a cytoskeletonlike structure (38). These proteins, ...

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Section: Discussionmentioning

confidence: 54%

Section: Detection Of P65 In Triton X-100-insoluble Subfractionsmentioning

confidence: 99%

The proline-rich P65 protein of Mycoplasma pneumoniae is a component of the Triton X-100-insoluble fraction and exhibits size polymorphism in the strains M129 and FH

et al. 1995

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“…Previous studies by us (33)(34)(35)(36) and other workers (4,7,8,20,31) demonstrated the presence of one copy of three distinct RepMPs within the cytadhesin P1 operon of M. pneumoniae. Since protein P1 is a major adhesin that also stimulates a strong host immune response, sequence variations resulting from RepMP-mediated recombination could contribute to M. pneumoniae disease pathogenesis.…”

Section: Discussionmentioning

confidence: 73%

“…Proteins encoded by the latter gene are required for P1 function and along with P1 likely constitute a major adhesin complex (40,41). Two of the repetitive elements found in the P1 gene, RepMP2/3 and RepMP4, have been studied in depth (31,34,35). Analysis of the complete genome sequence of M. pneumoniae strain M129 revealed 10 copies of RepMP2/3 and 8 copies of RepMP4 distributed throughout the chromosome (16).…”

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Genomic Analysis Reveals Mycoplasma pneumoniae Repetitive Element 1-Mediated Recombination in a Clinical Isolate

2008

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Mycoplasmas are cell wall-less bacteria that evolved by drastic reduction of the genome size. Complete genome analysis of Mycoplasma pneumoniae revealed the presence of numerous copies of four distinct large M. pneumoniae repetitive elements (RepMPs). One copy each of RepMP2/3, RepMP4, and RepMP5 are localized within the P1 operon (MPN140 to MPN142 loci), and their involvement in sequence variation in adhesin P1 and adherence-related protein B/C has been documented. Here we analyzed a clinical strain of M. pneumoniae designated S1 isolated from a 1993 outbreak of respiratory infections in San Antonio, TX. Based on the type of RepMPs within the P1 operon, we classified clinical isolate S1 as type 2 with unique minor sequence variations. Hybridization with oligonucleotide arrays revealed sequence divergence in two previously unsuspected hypothetical genes (MPN137 and MPN138 loci). Closer inspection of this region revealed that the MPN137 and MPN138 loci harbored previously unrecognized unique RepMP1 sequences found only in M. pneumoniae. PCR and sequence analyses revealed a recombination event involving three RepMP1-containing genes that resulted in fusion of MPN137 and MPN138 reading frames and loss of all but a short fragment of another RepMP1-containing locus, MPN130. The multiple copies of unique RepMP1 elements spread throughout the chromosome could allow vast numbers of sequence variations in clinical strains. Comparisons of amino acid sequences showed the presence of leucine zipper motifs in MPN130 and MPN138 proteins in reference strain M129 and the absence of these motifs in the fused protein of S1. The presence of tandem leucine and other repeats points to possible regulatory functions of proteins encoded by RepMP1-containing genes.Mycoplasma pneumoniae is a well-known human bacterial pathogen of the respiratory tract. The majority of clinical manifestations of M. pneumoniae infections range from tracheobronchitis and pharyngitis to pronounced signs of pneumonia and extrapulmonary pathologies. M. pneumoniae was identified in the early 1960s as the cause of primary atypical pneumonia, and some of the earliest documented outbreaks occurred in the military and in nonmilitary settings such as educational institutions, hospitals, and summer camps. In general, M. pneumoniae is among the most common causes of respiratory infections in school-age children and adults, and 20 to 30% of all community-acquired pneumonia cases are directly linked to M. pneumoniae. M. pneumoniae infections also lead to a spectrum of complicated sequelae, including asthma and other airway maladies, arthritis, pericarditis, central nervous system disorders, including encephalomyelitis, and autoimmune-associated pathologies (2, 39).M. pneumoniae is among the smallest self-replicating prokaryotic pathogens and has a genome size of 816 kb; about 8% of the genome is composed of repetitive DNA elements (RepMPs) (16). The existence of a high number of RepMPs, along with the loss of many metabolic functions during the process of genome reducti...

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The proteome of the bacteriumMycoplasma pneumoniae: Comparing predicted open reading frames to identified gene products

2002

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An existing proteome map of the bacterium Mycoplasma pneumoniae comprising proteins from 224 genes was extended to 305 genes. This corresponds to about 44% of the 688 proposed genome sequence derived open reading frames (ORFs). The newly assigned gene products were enriched, separated by one-dimensional or two-dimensional (2-D) gel electrophoresis and identified by mass spectrometry. The enrichment procedures included differential centrifugation, anion and cation exchange chromatography, affinity chromatography with heparin as a ligand and isolation of biotinylated proteins by binding to immobilized streptavidin. A comparative analysis of the identified proteins from 305 genes with the as yet unverified 383 ORFs concerning isoelectric point, molecular weight and number of transmembrane segments revealed that proteins with more than three predicted transmembrane segments and an isoelectric point above 10.5 are most likely not to be separated by 2-D gel electrophoresis. The mutual benefits of genomics and proteomics were shown by the identification of a todate unannotated 128 amino acid long protein.

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Analysis of three different repeated DNA elements present in the P1 operon ofMycoplasma pneumoniae: size, number and distribution on the genome

Cited by 69 publications

References 23 publications

The proline-rich P65 protein of Mycoplasma pneumoniae is a component of the Triton X-100-insoluble fraction and exhibits size polymorphism in the strains M129 and FH

The proline-rich P65 protein of Mycoplasma pneumoniae is a component of the Triton X-100-insoluble fraction and exhibits size polymorphism in the strains M129 and FH

Genomic Analysis Reveals Mycoplasma pneumoniae Repetitive Element 1-Mediated Recombination in a Clinical Isolate

The proteome of the bacteriumMycoplasma pneumoniae: Comparing predicted open reading frames to identified gene products

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