Cloning, expression and significance of MPT53 for identification of secreted proteins of Mycobacterium tuberculosis

Harald G. Wiker, Stephen Ll Michell, R. Glyn Hewinson, Eric Spierings, Sadamu Nagai, Morten Harboe*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

26 Citations (SciVal)

Abstract

Based on our N-terminal amino acid sequence of MPT53 and a deduced DNA sequence, we searched for the corresponding gene in the Mycobacterium tuberculosis genomic sequence at the Sanger centre, localizing mpt53 close to mpt70 and mpt83. The gene was cloned and expressed, followed by purification of MPT53 to homogeneity from recombinant M. smegmatis culture fluid. In MPT53 there is 60% identity with the active site of thioredoxin of M. tuberculosis (MPT46) with two cysteins in a CXXC motif, but MPT53 could not serve as an alternative substrate for thioredoxin reductase. Testing for IgM and IgG1 anti-MPT53 in cattle sera showed that MPT53 is immunogenic following natural and experimental infection with M. bovis. Cloning of mpt53 represents cloning of the last of the 10 proteins originally defined as 'secreted proteins' of M. tuberculosis and M. bovis based on determination of their 'Localization index' (LI). The need for a precise definition of the term 'secreted protein' is discussed. So far we have observed full concordance between occurrence of an LI value indicating secretion of a protein and occurrence of a signal sequence in the corresponding gene. Signal sequence independent protein secretion in mycobacteria may occur for a limited number of proteins and remains to be established.

Original languageEnglish
Pages (from-to)207-219
Number of pages13
JournalMicrobial Pathogenesis
Volume26
Issue number4
DOIs
Publication statusPublished - 01 Apr 1999

Keywords

  • MPT53
  • Mycobacterium bovis
  • Mycobacterium tuberculosis
  • Secreted proteins
  • Thioredoxin
  • Consensus Sequence
  • Chromatography, Ion Exchange
  • Mycobacterium bovis/chemistry
  • Mycobacterium tuberculosis/chemistry
  • Molecular Sequence Data
  • Thioredoxin-Disulfide Reductase/chemistry
  • Recombinant Proteins/chemistry
  • Cattle
  • Cloning, Molecular
  • Bacterial Proteins/chemistry
  • Amino Acid Sequence
  • Enzyme-Linked Immunosorbent Assay
  • Electrophoresis, Polyacrylamide Gel
  • Antigens, Bacterial
  • Antibodies, Monoclonal
  • Blotting, Western
  • Sequence Homology, Amino Acid
  • Sequence Alignment
  • Animals
  • Sequence Analysis
  • Gene Expression Regulation, Bacterial

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