ABSTRACT
BACKGROUND: Small heat shock proteins are ubiquitous family of stress proteins, having a role in virulence and survival of the pathogen. M. leprae, the causative agent of leprosy is an uncultivable organism in defined media, hence the biology and function of proteins were examined by cloning M. leprae genes in heterologous hosts. The study on sHsp18 was carried out as the knowledge about the functions of this major immunodominant antigen of M. leprae is scanty. RESULTS: The gene encoding Mycobacterium leprae small heat shock protein (sHsp18) was amplified from biopsy material of leprosy patients, and cloned and expressed in E. coli. The localization and in vitro characterization of the protein are detailed in this report. Data show that major portion of the protein is localized in the outer membrane of E. coli. The purified sHsp18 functions as an efficient chaperone as shown by their ability to prevent thermal inactivation of restriction enzymes SmaI and NdeI. Physical interaction of the chaperone with target protein is also demonstrated. Size exclusion chromatography of purified protein shows that the protein can form multimeric complexes under in vitro conditions as is demonstrated for several small heat shock proteins. CONCLUSION: The small heat shock protein sHsp18 of M. leprae is a chaperone and shows several properties associated with other small heat shock proteins. Membrane association and in vitro chaperone function of sHsp18 shows that the protein may play a role in the virulence and survival of M. leprae in infected host.
Subject(s)
Bacterial Proteins/metabolism , Heat-Shock Proteins, Small/metabolism , Leprosy/microbiology , Mycobacterium leprae/metabolism , Bacterial Outer Membrane Proteins/chemistry , Bacterial Outer Membrane Proteins/genetics , Bacterial Outer Membrane Proteins/metabolism , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Escherichia coli/genetics , Escherichia coli/metabolism , Gene Expression , Heat-Shock Proteins, Small/chemistry , Heat-Shock Proteins, Small/genetics , Humans , Molecular Chaperones/chemistry , Molecular Chaperones/genetics , Molecular Chaperones/metabolism , Mycobacterium leprae/chemistry , Mycobacterium leprae/genetics , Protein Binding , Protein TransportABSTRACT
In this work we are proposing Homology modeled structures of Mycobacterium leprae 18kDa heat shock protein and its mutant. The more closely related structure of the small heat shock protein (sHSP) belonging to the eukaryotic species from wheat sHSP16.9 and 16.3kDa ACR1 protein from Mycobacterium tuberculosis were used as template structures. Each model contains an N-terminal domain, alpha-crystalline domain and a C-terminal tail. The models showed that a single point mutation from serine to proline at 52(nd) position causes structural changes. The structural changes are observed in N-terminal region and alpha-crystalline domains. Serine in 52(nd) position is observed in beta4 strand and Proline in 52(nd) position is observed in loop. The number of residues contributing alpha helix at N-terminal region varies in both models. In 18S more number of residues is present in alpha helix when compared to 18P. The loop regions between beta3 and beta4 strands of both models vary in number of residues present in it. Number of residues contributing beta4 strand in both models vary. beta6 strand is absent in both models. Major functional peptide region of alpha crystalline domains of both models varies. These differences observed in secondary structures support their distinct functional roles. It also emphasizes that a point mutation can cause structural variation.
