Expression and immunologic characterization of recombinant heat shock protein 58 of Leptospira species: A major target antigen of the humoral immune response

Se Hoon Park, Byung-Yoon Ahn, Ja Kim Min

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Abstract

A clone of Leptospira interrogans serovar lai that was isolated by immunoscreening of a genomic λ library with sera from convalescent patients with leptospirosis directed expression of a unique 62-kDa protein in Escherichia coli. When examined by SDS-PAGE, the protein comigrated with an immunodominant protein present in leptospiral cell lysate. Determination of the nucleotide sequence of the 2.7-kb insert DNA identified two genes homologous to the hsp58 and hsp10 of L. interrogans serovar copenhageni reported previously. The overexpressed recombinant Hsp58 protein was purified and used to immunize a rabbit to produce a polyclonal antibody. Immunoblot analysis using the rabbit anti-Hsp58 G antibodies showed that the 62-kDa protein was commonly present in lysates of other serovars of leptospires, consistent with the strong sequence conservation between the hsp58 genes of the two serovars. Immunoglobulin G antibodies to the Hsp58 were specifically detected by ELISA in 82% of sera (18/22) from patients with leptospirosis. Deletion analysis of the recombinant Hsp58 protein indicated that a strong antigenic determinant for humoral immune response is located between amino acids 360 and 380 (DREKLQERLAKLAGGVAVIHV) of Hsp58, which are highly conserved among the GroEL family. The strong sequence conservation of the Hsp58 among leptospires and its importance as a major target for the humoral immune response warrant further studies of its potential pathogenetic role.

Original languageEnglish
Pages (from-to)903-910
Number of pages8
JournalDNA and Cell Biology
Volume18
Issue number12
DOIs
Publication statusPublished - 1999 Dec 1

Fingerprint

Leptospira
Humoral Immunity
Heat-Shock Proteins
Recombinant Proteins
Leptospira interrogans
Antigens
Leptospirosis
Antibodies
Rabbits
Proteins
Genomic Library
Escherichia coli Proteins
Serum
Genes
Epitopes
Polyacrylamide Gel Electrophoresis
Clone Cells
Immunoglobulin G
Enzyme-Linked Immunosorbent Assay
Amino Acids

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cell Biology

Cite this

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abstract = "A clone of Leptospira interrogans serovar lai that was isolated by immunoscreening of a genomic λ library with sera from convalescent patients with leptospirosis directed expression of a unique 62-kDa protein in Escherichia coli. When examined by SDS-PAGE, the protein comigrated with an immunodominant protein present in leptospiral cell lysate. Determination of the nucleotide sequence of the 2.7-kb insert DNA identified two genes homologous to the hsp58 and hsp10 of L. interrogans serovar copenhageni reported previously. The overexpressed recombinant Hsp58 protein was purified and used to immunize a rabbit to produce a polyclonal antibody. Immunoblot analysis using the rabbit anti-Hsp58 G antibodies showed that the 62-kDa protein was commonly present in lysates of other serovars of leptospires, consistent with the strong sequence conservation between the hsp58 genes of the two serovars. Immunoglobulin G antibodies to the Hsp58 were specifically detected by ELISA in 82{\%} of sera (18/22) from patients with leptospirosis. Deletion analysis of the recombinant Hsp58 protein indicated that a strong antigenic determinant for humoral immune response is located between amino acids 360 and 380 (DREKLQERLAKLAGGVAVIHV) of Hsp58, which are highly conserved among the GroEL family. The strong sequence conservation of the Hsp58 among leptospires and its importance as a major target for the humoral immune response warrant further studies of its potential pathogenetic role.",
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