Characterization of the gene encoding the immunodominant 35 kDa protein of Mycobacterium leprae

Autor: Maria Cristina Vidal Pessolani, Nathalie Winter, Patrick J. Brennan, Karin Eiglmeier, Becky Rivoire, Warwick J. Britton, James A. Triccas, Eng-Mong Lim, S W Hunter
Přispěvatelé: Centenary Institute of Cancer Medicine and Cell Biology, Colorado State University [Fort Collins] (CSU), Institut Pasteur [Paris], Génétique mycobactérienne - Mycobacterial genetics, Institut Pasteur [Paris] (IP)
Jazyk: angličtina
Rok vydání: 1995
Předmět:
T-Lymphocytes
Restriction Mapping
Polymerase Chain Reaction
Promoter Regions
Genetic
Mycobacterium leprae
Genes
Dominant
0303 health sciences
biology
Mycobacterium smegmatis
Antibodies
Monoclonal
Cosmids
Recombinant Proteins
3. Good health
BOVIS BCG
[SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology
Mycobacterium tuberculosis complex
Mycobacterium fortuitum
EXPRESSION
Protein subunit
Molecular Sequence Data
LEPROSY
TUBERCULOSIS
Microbiology
SEQUENCE
ANTIGENS
Mycobacterium
Gene product
CLONING
03 medical and health sciences
Bacterial Proteins
HSPA2
Humans
Amino Acid Sequence
Molecular Biology
030304 developmental biology
DNA Primers
Gene Library
Antigens
Bacterial
Base Sequence
Sequence Homology
Amino Acid
030306 microbiology
STRAINS
Gene Expression Regulation
Bacterial
biology.organism_classification
Molecular biology
PERFORMANCE LIQUID-CHROMATOGRAPHY
Molecular Weight
Genes
Bacterial
MONOCLONAL-ANTIBODIES
Zdroj: Molecular Microbiology
Molecular Microbiology, Wiley, 1995, 16 (5), pp.865-876. ⟨10.1111/j.1365-2958.1995.tb02314.x⟩
Molecular Microbiology, 1995, 16 (5), pp.865-876. ⟨10.1111/j.1365-2958.1995.tb02314.x⟩
ISSN: 0950-382X
1365-2958
DOI: 10.1111/j.1365-2958.1995.tb02314.x⟩
Popis: International audience; Analysis of the interaction between the host immune system and the intracellular parasite Mycobacterium leprae has identified a 35 kDa protein as a dominant antigen, The native 35 kDa protein was purified from the membrane fraction of M. leprae and termed MMPI (major membrane protein I). As the purified protein was not amenable to N-terminal sequencing, partial proteolysis was used to establish the sequences of 21 peptides, A fragment of the 35 kDa proteinen-coding gene was amplified by the polymerase chain reaction from M. leprae chromosomal DNA with oligonucleotide primers derived from internal peptide sequences and the whole gene was subsequently isolated from a M. leprae cosmid library. The nucleotide sequence of the gene revealed an open reading frame of 307 amino acids containing most of the peptide sequences derived from the native 35 kDa protein, The calculated subunit mass was 33.7 kDa, but the native protein exists as a multimer of 950 kDa, Database searches revealed no identity between the 35 kDa antigen and known protein sequences. The gene was expressed in Mycobacterium smegmatis under the control of its own promoter or at a higher level using an 'up-regulated' promoter derived from Mycobacterium fortuitum. The gene product reacted with monoclonal antibodies raised to the native protein, Using the bacterial alkaline phosphatase reporter system, we observed that the 35 kDa protein was unable to be exported across the membrane of recombinant M. smegmatis. The 35 kDa protein-encoding gene is absent from members of the Mycobacterium tuberculosis complex, but homologous sequences were detected in Mycobacterium avium, Mycobacterium haemophilum and M. smegmatis, The avaibility of the recombinant 35 kDa protein will permit dissection of both antibody- and T-cell-mediated immune responses in leprosy patients.
Databáze: OpenAIRE
Externí odkaz: