Selection and characterization of recombinant clones that produce Mycobacterium leprae antigens recognized by antibodies in sera from household contacts of leprosy patients

Hartskeerl, Rudy A.; Rens, R. M. Van; Stabel, L F; Wit, M. Y. L. De; Klatser, P. R.

doi:10.1128/iai.58.9.2821-2827.1990

Cited by 11 publications

(4 citation statements)

References 36 publications

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“…Nothing is known about the possible presence of an nrdAB-like operon in M. tuberculosis. Moreover, the sequence of a fragment of an nrdA-like gene of Mycobacterium leprae (corresponding to residues 433-482 of its homologous R1 protein of S. typhimurium ) has been determined and investigated for its significant identity with the human R1 (Hartskeerl et al, 1990). The identity between this nrdA fragment of M. leprae and S. typhimurium is dramatically lower (27%) than that shown by the M. tuberculosis and S. typhimurium nrdE genes.…”

Section: Discussionmentioning

confidence: 99%

Promoter identification and expression analysis of Salmonella typhimurium and Escherichia coli nrdEF operons encoding one of two class I ribonucleotide reductases present in both bacteria

Jordan

Aragall

Gibert

et al. 1996

Molecular Microbiology

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Salmonella typhimurium and Escherichia coli cells have two different class I ribonucleotide reductases encoded by the nrdEF and nrdAB operons. Despite the presence of one additional ribonucleotide reductase, the nrdAB-encoded enzyme is essential to the aerobic growth of the cell because nrdAB-defective mutants of both species are not viable in the presence of oxygen. Several factors controlling nrdAB gene transcription have been analysed intensively. Nothing is known about the expression of the nrdEF genes. To study this subject, and after cloning of E. coli nrdEF genes and sequencing of their 5' ends, the promoter of this operon has been identified by primer extension in both bacterial species. The +1 position was 691 bp and 692 bp upstream of the translational start points of the nrdE genes of S. typhimurium and E. coli, respectively. Downstream of the +1 position, and before the nrdE gene, two open reading frames (ORFs) of 81 and 136 amino acid residues are present in both bacteria. The synthesis of a polypeptide with a molecular mass of 9 kDa, corresponding to the first of these two ORFs, was observed by using the T7 RNA polymerase expression system. Comparison of the amino acid predicted sequence of this ORF reveals a significant similarity with glutaredoxin proteins. Competitive, reverse-transcription polymerase chain reaction experiments indicate that transcription from the nrdEF promoter normally takes place in wild-type cells. nrdEF transcription is increased by hydroxyurea, which inhibits class I ribonucleotide reductase activity, in both RecA+ and RecA- cells. nrdA(ts) mutants show a higher level of nrdEF transcription than wild-type cells at either the permissive or the restrictive temperature. nrdEF expression was unaffected by changes in DNA supercoiling whether caused by the introduction of either topA::Tn10 and hns::Tn10 mutations or by the inhibition of DNA gyrase with the antibiotic novobiocin. In contrast to the nrdAB genes, the nrdEF operon is not essential to the cells because nrdEF-defective mutants are viable under both aerobic and anaerobic conditions.

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Section: Discussionmentioning

confidence: 99%

Promoter identification and expression analysis of Salmonella typhimurium and Escherichia coli nrdEF operons encoding one of two class I ribonucleotide reductases present in both bacteria

Jordan

Aragall

Gibert

et al. 1996

Molecular Microbiology

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“…The construction of a Mycobacterium leprae gene expression library in lambda gti 1 has been a major tool for isolating M. leprae protein antigens by screening with either monoclonal antibodies (mAbs) against M. teprae (Young et ai, 1985) or with sera of leprosy patients (e.g. Sathish ef at., 1990;Hartskeerl et at.. 1990;Cherayil and Young, 1988). Among the antigens that were isolated, the 65 kDa antigen appeared to be immunodominant in both the B-cell and T-cell responses against the M. teprae bacillus (Thole and van der Zee, 1990).…”

Section: Introductionmentioning

confidence: 99%

Mycobacteria contain two groEL genes: the second Mycobacterium leprae groEL gene is arranged in an operon with groES

Wit

Bekelie

Osland

et al. 1992

Molecular Microbiology

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In contrast to other bacterial species, mycobacteria were thus far considered to contain groEL and groES genes that are present on separate loci on their chromosomes, Here, by screening a Mycobacterium leprae lambda gt11 expression library with serum from an Ethiopian lepromatous leprosy patient, two DNA clones were isolated that contain a groEL gene arranged in an operon with a groES gene. The complete DNA sequence of this groESL operon was determined. The predicted amino acid sequences of the GroES and GroEL proteins encoded by this operon are 85-90% and 59-61% homologous to the sequences from previously characterized mycobacterial GroES and GroEL proteins. Southern blotting analyses with M. leprae groES- and groEL-specific probes demonstrate that similar groESL homologous DNA is present in the genomes of other mycobacteria, including Mycobacterium tuberculosis. This strongly suggests that mycobacteria contain a groESL operon in addition to a separately arranged second groEL gene. Using five T-cell clones from two leprosy patients as probes, expression of the M. leprae GroES protein in Escherichia coli after heat shock was demonstrated. Four of these clones recognized the same M. leprae-specific GroES-derived peptide in a DR2-restricted fashion. No expression of the groEL gene from this operon was detected in E. coli after heat shock, as tested with a panel of T-cell clones and monoclonal antibodies reactive to previously described GroEL proteins of mycobacteria.

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“…None of the antigens studied to date has emerged as a suitable candidate for development of a diagnostic assay for early tuberculosis. Since antigens and epitopes recognized during natural infection and disease progression in humans may differ substantially from those recognized by animals upon artificial immunization (5,8,26,33,35,47), a selection of antigens based on their ability to stimulate the human immune system is required.…”

mentioning

confidence: 99%

Human humoral responses to antigens of Mycobacterium tuberculosis: immunodominance of high-molecular-mass antigens

Laal

Samanich

Sonnenberg

et al. 1997

Clin Diagn Lab Immunol

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The selection of antigens of Mycobacterium tuberculosis for most studies of humoral responses in tuberculosis patients has been restricted to molecules that were either immunodominant in immunized animals or amenable to biochemical purification rather than those that were reactive with the human immune system. Delineation of antigens that elicit humoral responses during the natural course of disease progression in humans has been hindered by the presence of cross-reactive antibodies to conserved regions on ubiquitous prokaryotic antigens in sera from healthy individuals and tuberculosis patients. The levels of cross-reactive antibodies in the sera were reduced by preadsorption with Escherichia coli lysates, prior to studying their reactivity against a large panel of M. tuberculosis antigens to which the human immune system may be exposed during natural infection and disease. Thus, reactivity against pools of secreted, cellular, and cell wallassociated antigens of M. tuberculosis was assessed by an enzyme-linked immunosorbent assay (ELISA). Initial results suggested that the secreted protein preparation contained antigens most frequently recognized by the humoral responses of pulmonary tuberculosis patients. The culture filtrate proteins were subsequently size fractionated by preparative polyacrylamide gel electrophoresis, characterized by reaction with murine monoclonal antibodies to known antigens of M. tuberculosis by an ELISA, and assessed for reactivity with tuberculous and nontuberculous sera. Results show that a secreted antigen of 88 kDa elicits a strong antibody response in a high percentage of patients with pulmonary tuberculosis. This and other antigens identified on the basis of their reactivity with patient sera may prove useful for developing serodiagnosis for tuberculosis.

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Selection and characterization of recombinant clones that produce Mycobacterium leprae antigens recognized by antibodies in sera from household contacts of leprosy patients

Cited by 11 publications

References 36 publications

Promoter identification and expression analysis of Salmonella typhimurium and Escherichia coli nrdEF operons encoding one of two class I ribonucleotide reductases present in both bacteria

Promoter identification and expression analysis of Salmonella typhimurium and Escherichia coli nrdEF operons encoding one of two class I ribonucleotide reductases present in both bacteria

Mycobacteria contain two groEL genes: the second Mycobacterium leprae groEL gene is arranged in an operon with groES

Human humoral responses to antigens of Mycobacterium tuberculosis: immunodominance of high-molecular-mass antigens

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