Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0026918 (Mycobacterium)
 52,428 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

With the advent of the sequencing programs of prokaryotic genomes, many examples of the presence of serine/threonine protein kinases in these organisms have been identified. Moreover, these kinases could be classified as homologues of those belonging to the well characterized superfamily of the eukaryotic serine/threonine and tyrosine kinases. Eleven such kinases were recognized in the genome of Mycobacterium tuberculosis. Here we report the crystal structure of an active form of PknB, one of the four M. tuberculosis kinases that are conserved in the downsized genome of Mycobacterium leprae and are therefore presumed to play an important role in the processes that regulate the complex life cycle of mycobacteria. Our structure confirms again the extraordinary conservation of the protein kinase fold and constitutes a landmark that extends this conservation across the evolutionary distance between high eukaryotes and eubacteria. The structure of PknB, in complex with a nucleotide triphosphate analog, reveals an enzyme in the active state with an unprecedented arrangement of the Gly-rich loop associated with a new conformation of the nucleotide gamma-phosphoryl group. It presents as well a partially disordered activation loop, suggesting an induced fit mode of binding for the so far unknown substrates of this kinase or for some modulating factor(s).
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PMID:Crystal structure of the catalytic domain of the PknB serine/threonine kinase from Mycobacterium tuberculosis. 1255 95

The enzyme hydroxymethylbilane synthase (HMBS, EC 4.3.1.8), 313 amino acid residues and MW 34 kDa, also known as porphobilinogen deaminase (PBGD), catalyses the stepwise polymerization of four molecules of porphobilinogen (PBG) to the linear tetrapyrrole 1-hydroxymethylbilane. Several crystallographic structures of HMBS have been previously determined, most recently including by time-resolved Laue protein crystallography of the Lys59Gln mutant form with reaction initiation undertaken by use of a flow cell carrying the substrate PBG. In this paper we review these structures and add new molecular graphics representations and analyses. Moreover we present a new structure refined at 1.66 A resolution using diffraction data recorded at cryo-temperature (100 K) in an attempt at trapping the polypeptide loop (residues 47 to 58) in the vicinity of the enzyme active site, missing in all previous structure determinations. This loop still has not appeared in the electron density maps, in spite of the advantage of cryo-temperature, but nevertheless the 1.66 A cryo-structure extends the ensemble of known HMBS structures. The cryomodel of protein, cofactor and 320 bound water molecules has been refined to a final R-factor and R-free of 0.198 and 0.247 respectively; the PDB deposition codes, coordinates and structure factors are 1GTK and R1GTKSF respectively. Finally a protein comparison study is presented of the Mycobacterium tuberculosis (MTb) HMBS, with the E. coli HMBS. This has been done as preparation for future structural studies on the MTb HMBS from this important disease bearing organism. The overall amino acid sequence identity is 41%. Most interestingly there is a two-residue reduction in length of the loop referred to above (Asp 50 and Gly 58 being missing in the MTb form). This gives the hope that this loop will be less flexible and thus might become visible to crystallographic analysis.
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PMID:Time-resolved and static-ensemble structural chemistry of hydroxymethylbilane synthase. 1255 54

The katG, inhA and ahpC genes, in 71 isoniazid (INH)-resistant and 26 INH-susceptible Mycobacterium tuberculosis isolates, from South Korea were examined by sequencing and MspI restriction enzyme analysis. Mutations in the katG 315 alone, katG 315 plus inhA, katG 315 plus ahpC, katG 309 alone, katG 309 plus inhA, inhA alone, and ahpC alone, were detected in 54.9, 2.8, 1.4, 1.4, 1.4, 19.7, and 5.6% of the 71 INH-resistant isolates, respectively. There was no statistically significant difference (p > 0.05) in the frequencies of these mutations for the INH-monoresistant compared with the multidrug-resistant isolates. Mutations in the katG codon 315 were associated with the high-level of INH resistance (MIC, >1 microg/ml), whereas the mutation in the inhA promoter region was associated with the low-level of INH resistance (MIC, >0.2 to 1 microg/ml). The previously undescribed GGT-->GAT (Gly-->Asp) mutation in the katG codon 309 was found in two rifampin, including-multidrug-resistant isolates, but we cannot assess if this is predictive of INH resistance. The sensitivity and specificity of molecular analysis of the katG codon 315 and/or the inhA promoter region were 80.3 and 100%, respectively. Therefore, mutations in these regions are highly predictive of INH resistance in South Korea.
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PMID:Molecular analysis of isoniazid resistance in Mycobacterium tuberculosis isolates recovered from South Korea. 1459 68

Continuing our research on Mycobacteria kansasii phagocytosis inhibition, we have examined in that context three series of peptides derived from the RGDVY and GRGD sequences. It was found that the levels of the inhibitory activity depend on the amino acid composition as well as on the particular peptide sequence. Distinct inhibitory activity was found in the case of thymopentin (RKDVY), the active fragment of thymopoietin. In this case the Mycobacterium phagocytosis inhibition should be combined with general immunostimulatory activity of RKDVY peptide. Our examination of a series of GRGDV analogs with a successively prolonged oligo-Gly linker inserted into the peptide chain showed that the distance between the Arg and Asp residues required for such an activity should be about 9A.
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PMID:Analogs of RGDVY and GRGD peptides inhibit Mycobacterium kansasii phagocytosis. 1461 80

Mycobacterium tuberculosis is able to persist in the human host for decades in an apparently dormant state where it is presumed to reside in an hypoxic environment. This can be mimicked by the Wayne culture model in which progressive oxygen depletion causes the bacteria to shift into a non-replicating state. We investigated global gene expression in aerobic (roller), microaerophilic (NRP1) and anaerobic (NRP2) cultures. A number of genes were significantly up-regulated as compared to aerobic culture; 178 in NRP1, 210 in NRP2, 88 in both. The two states showed distinct gene expression profiles, although a number of membrane and transmembrane proteins were induced in both conditions. A number of regulatory proteins were up-regulated in NRP2. Glycine dehydrogenase, nitrate reductase and alpha-crystallin were induced in both stages, as were fatty acid metabolism genes including fadD26 and mas and genes of the DosR regulon. In a comparison with other stress conditions, there were more similarities between anaerobic conditions and carbon starvation or heat shock than between microaerophilic conditions and carbon starvation or heat shock, but as expected microaerophilic and anaerobic conditions showed the most similar profile. Our results indicate that a large number of genes are up-regulated during the shift into the persistent state.
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PMID:Gene expression profile of Mycobacterium tuberculosis in a non-replicating state. 1520 93

The function of the PE/PPE families of proteins, which represent approximately 10% of the coding capacity of the Mycobacterium tuberculosis genome, has remained relatively unknown. We earlier described a PPE family member, Rv2430c, as an immunodominant antigen. We now report another PPE family gene, Rv2608, a member of the major polymorphic tandem repeat subfamily, for its ability to elicit a high humoral and a low T cell response. Rv2608 was also found to be polymorphic in different clinical isolates of M. tuberculosis, as determined by polymerase chain reaction-restriction fragment-length polymorphism analysis. A total of 51 clinically confirmed patients with tuberculosis (TB), belonging to 3 different categories--fresh infection (n=22), relapsed infection (n=21), and extrapulmonary infection (n=8)--and 10 healthy control subjects were included in the study. Recombinant Rv2608 protein showed positive reactivity to patients' serum samples. Enzyme-linked immunosorbent assays and T cell-proliferation assays with synthetic peptides corresponding to predicted regions of high antigenicity showed a predominantly humoral response in patients with relapsed TB. We additionally identified the Gly-X-Gly-Asn-X-Gly repeat motifs as being primarily responsible for eliciting a humoral immune response.
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PMID:Regions of high antigenicity within the hypothetical PPE major polymorphic tandem repeat open-reading frame, Rv2608, show a differential humoral response and a low T cell response in various categories of patients with tuberculosis. 1534 33

Mycobacterium tuberculosis KatG enzyme functions both as catalase for removing hydrogen peroxide (H(2)O(2)) and as peroxidase for oxidating isoniazid (INH) to active form of anti-tuberculosis drug. Although mutations in M. tuberculosis KatG confer INH resistance in tuberculous patients, structural bases for INH-resistant mutations in the KatG gene remains poorly understood. Here, three M. tuberculosis KatG mutants bearing Arg418--> Gln, Ser315 --> Thr, or Trp321 --> Gly replacement were assessed for changes in catalase-peroxidase activities and possible structure bases relevant to such changes. These three M. tuberculosis KatG mutants exhibited a marked impairment or loss of catalase-peroxidase activities. The possible structural bases for the mutant-induced loss of enzyme activities were then analyzed using a three-dimensional model of M. tuberculosis KatG protein constructed on the basis of the crystal structure of the catalase-peroxidase from Burkholderia pseudomallei. The model suggests that three M. tuberculosis KatG mutants bearing Arg418 --> Gln, Ser315 -->Thr, or Trp321--> Gly replacement affect enzyme activities by different mechanisms, although each of them impacts consequently on a heme-associated structure, the putative oxidative site. Moreover, in addition to the widely accepted substrate-binding site, M. tuberculosis KatG may bear another H(2)O(2) binding site. This H(2)O(2) binding site appears to interact with the catalytic site by a possible electron-transfer chain, a Met255-Tyr229-Trp107 triad conserved in many catalase-peroxidases. The Ser315 --> Thr mutant may have direct effect on the catalytic site by interfering with electron transfer in addition to the previously proposed mechanism of steric constraint.
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PMID:Three-dimensional model and molecular mechanism of Mycobacterium tuberculosis catalase-peroxidase (KatG) and isoniazid-resistant KatG mutants. 1565 Mar 70

Three MutS fusion systems were designed as the mutation recognition and signal elements of DNA chips for detection of DNA mutations. The expression vectors containing the encoding sequences of three recombinant proteins, Trx-His6-GFP-(Ser-Gly)6-MutS (THGLM), Trx-His6-(Ser-Gly)6-Strep tagII-(Ser-Gly)6-MutS (THLSLM) and Trx-His6-(Ser-Gly)6-MutS (THLM), were constructed by gene slicing in vitro. THGLM, THLSLM and THLM were then expressed in Escherichia coli AD494(DE3), respectively. SDS-PAGE analysis revealed that each of the expected proteins was approximately 30% of the total bacterial proteins. The recombinant proteins were purified to the purity over 90% by immobilized metal (Co2+) chelation affinity chromatography. Bioactivity assay indicated that three fusion proteins retained the mismatch-binding activity and the functions of other fusion partners. DNA chips arrayed both mismatched and unpaired DNA oligonucleotides as well as rpoB gene from Mycobacterium tuberculosis were prepared. THGLM, THLSLM and THLM that was labeled with Fluorolinktrade mark Cy3 reactive dye, were then used as both mutation recognition and labeling elements of DNA chips. The resulting DNA chips were used to detect the mismatched and unpaired mutations in the synthesized oligonucleotides and single base mutation in rpoB gene of M. tuberculosis that is resistant to rifamycin.
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PMID:Construction and characterization of different MutS fusion proteins as recognition elements of DNA chip for detection of DNA mutations. 1596 61

Proteins of the 6-kDa early secreted antigenic target (ESAT-6) secretion system-1 of Mycobacterium tuberculosis are not only strongly involved in the anti-mycobacterial Th1-host immune response but are also key players for virulence. In this study, protein engineering together with bioinformatic, immunological, and virulence analyses allowed us to pinpoint regions of the ESAT-6 molecule that are critical for its biological activity in M. tuberculosis. Mutation of the Trp-Xaa-Gly motif, conserved in a wide variety of ESAT-6-like proteins, abolished complex formation with the partner protein CFP-10, induction of specific T-cell responses, and virulence. Replacement of conserved Leu residues interfered with secretion, coiled-coil formation, and virulence, whereas certain mutations at the extreme C terminus did not affect secretion but caused attenuation, possibly because of altered ESAT-6 targeting or trafficking. In contrast, the mutation of several residues on the outer surface of the four-helical bundle structure of the ESAT-6.CFP-10 complex showed much less effect. Construction of recombinant BCG expressing ESAT-6 with a C-terminal hexahistidine tag allowed us to co-purify ESAT-6 and CFP-10, experimentally confirming their strong interaction both in and outside of the mycobacterial cell. The strain induced potent, antigen-specific T-cell responses and intermediate in vivo growth in mice, suggesting that it remained immunogenic and biologically active despite the tag. Together with previous NMR data, the results of this study have allowed a biologically relevant model of the ESAT-6.CFP-10 complex to be constructed that is critical for understanding the structure-function relationship in tuberculosis pathogenesis.
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PMID:Functional analysis of early secreted antigenic target-6, the dominant T-cell antigen of Mycobacterium tuberculosis, reveals key residues involved in secretion, complex formation, virulence, and immunogenicity. 1604 98

Among fluoroquinolone-resistant Mycobacterium tuberculosis (FQr-MTB) isolates, mutation at positions 90, 91, and 94 in gyrA gene and at positions 495, 516, and 533 in gyrB gene have been frequently reported. In this study, 35 isolates of FQr-MTB were collected from Siriraj Hospital and Chest Disease Institute. The quinolone-resistance-determining regions (QRDR) of gyrA and gyrB genes in all 35 FQr-MTB isolates and from the H37Ra MTB strain were amplified using polymerase chain reaction (PCR). DNA-sequencing and single-strand conformation polymorphism (SSCP) were further utilized for characterization of the mutations in the QRDR of gyrA and gyrB genes and mutation screening, respectively. From DNA-sequencing, 21 of 35 (60%) exhibited single-point mutations in different positions, at Ala90Val, Ser91Pro, and Asp94(Gly/Ala/His/Asn); and one novel mutation position at Gly88Cys in the gyrA gene and Asp495Asn in the gyrB gene. These positions were previously frequently reported to be responsible for FQr-MTB. The other 14 FQr-MTB isolates (40%) had no mutation. This study is the first report of mutation occurring only in the QRDR of the gyrB gene, without prior mutation in the gyrA QRDR among FQr-MTB isolates. By SSCP analysis for screening of the mutant FQr-MTB, the SSCP patterns of mutated FQr-MTB isolates were clearly differentiated from the SSCP patterns of FQs-MTB.
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PMID:Mutations in the gyrA and gyrB genes of fluoroquinolone-resistant Mycobacterium tuberculosis from TB patients in Thailand. 1643 15


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