Immunological characterization of a bacterial protein isolated from salmonid fish naturally infected with Piscirickettsia salmonis.
The Salmon Rickettsia syndrome (SRS) remains a major infectious disease in the Chilean aquaculture. A limited number of Piscirickettsia salmonis proteins have been characterized so far for their use as potential candidates for vaccines studies. In this study, we identified and expressed a highly immunogenic protein of P. salmonis extracted by selective hydrophobicity from crude-cell macerates of naturally infected salmonid fish. One and two-D PAGE gels followed by Western blot analysis with a battery of polyclonal anti-P. salmonis antibodies have allowed the isolation of the target protein. Basic local alignment search (BLAST) done after partial sequencing of the pure protein identified it as a member of the heat-shock protein (HSP) family of prokaryotes. The protein, named ChaPs, was cloned as a single open reading frame encoding 545 amino acid residues with a predicted molecular mass of 57.3 kDa. The amplicon representing the entire novel gene was expressed in vitro in different heterologous systems: the PurePro Caulobacter crescentus expression system from where most of the characterization was attained, and also in the Escherichia coli BL-21 CodonPlus model for commercially potential purposes. The immunologic potential of ChaPs was determined with serum from naturally infected fish.
Marshall
Sergio H
SH
Laboratorio de Genética e Inmunología Molecular, Instituto de Biología, Facultad de Ciencias Básicas y Matemáticas, Pontificia Universidad Católica de Valparaíso, Avenida Brasil 2950, Valparaíso, Chile. smarshal@ucv.cl
Conejeros
Pablo
P
Zahr
Marcela
M
Olivares
Jorge
J
Gómez
Fernando
F
Cataldo
Patricio
P
Henríquez
Vitalia
V
D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D015153:Blotting, Western; D016934:Caulobacter / Q000235:genetics; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005393:Fish Diseases / Q000276:immunology / Q000382:microbiology; D015870:Gene Expression; D006360:Heat-Shock Proteins / Q000276:immunology* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017684:Oncorhynchus kisutch; D053522:Piscirickettsia / Q000276:immunology*; D044225:Piscirickettsiaceae Infections / Q000276:immunology / Q000382:microbiology / Q000662:veterinary*; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D006360:Heat-Shock Proteins