Molecular characterization of a novel proto-type antimicrobial protein galectin-1 from striped murrel.
In this study, we reported a molecular characterization of a novel proto-type galectin-1 from the striped murrel Channa striatus (named as CsGal-1). The full length CsGal-1 was identified from an established striped murrel cDNA library and further we confirmed the sequence by cloning. The complete cDNA sequence of CsGal-1 is 590 base pairs (bp) in length and its coding region encoded a poly peptide of 135 amino acids. The polypeptide contains a galactoside binding lectin domain at 4-135. The domain carries a sugar binding site at 45-74 along with its signatures (H(45)-X-Asn(47)-X-Arg(49) and Trp(69)-X-X-Glu(72)-X-Arg(74)). CsGal-1 shares a highly conserved carbohydrate recognition domain (CRD) with galectin-1 from other proto-type galectin of teleosts. The mRNA expressions of CsGal-1 in healthy and various immune stimulants including Aphanomyces invadans, Aeromonas hydrophila, Escherchia coli lipopolysaccharide and poly I:C injected tissues of C. striatus were examined using qRT-PCR. CsGal-1 mRNA is highly expressed in kidney and is up-regulated with different immune stimulants at various time points. To understand its biological activity, the coding region of CsGal-1 gene was expressed in an E. coli BL21 (DE3) cloning system and its recombinant protein was purified. The recombinant CsGal-1 protein was agglutinated with mouse erythrocytes at a concentration of 4μg/mL in a calcium independent manner. CsGal-1 activity was inhibited by d-galactose at 25mM(-1) and d-glucose and d-fructose at 100mM(-1). The results of microbial binding assay showed that the recombinant CsGal-1 protein agglutinated only with the Gram-negative bacteria. Interestingly, we observed no agglutination against Gram-positive bacteria. Overall, the study showed that CsGal-1 is an important immune gene involved in the recognition and elimination of pathogens in C. striatus.
Division of Fisheries Biotechnology & Molecular Biology, Department of Biotechnology, Faculty of Science and Humanities, SRM University, Kattankulathur 603203, Chennai, Tamil Nadu, India; Department of Microbiology, SRM Arts & Science College, Kattankulathur 603203, Chennai, Tamil Nadu, India.
Abirami
A
Arasu
Division of Fisheries Biotechnology & Molecular Biology, Department of Biotechnology, Faculty of Science and Humanities, SRM University, Kattankulathur 603203, Chennai, Tamil Nadu, India.
Venkatesh
V
Kumaresan
Division of Fisheries Biotechnology & Molecular Biology, Department of Biotechnology, Faculty of Science and Humanities, SRM University, Kattankulathur 603203, Chennai, Tamil Nadu, India; Department of Biotechnology, SRM Arts & Science College, Kattankulathur 603203, Chennai, India.
Akila
A
Sathyamoorthi
Division of Fisheries Biotechnology & Molecular Biology, Department of Biotechnology, Faculty of Science and Humanities, SRM University, Kattankulathur 603203, Chennai, Tamil Nadu, India.
Mukesh Kumar
MK
Chaurasia
Division of Fisheries Biotechnology & Molecular Biology, Department of Biotechnology, Faculty of Science and Humanities, SRM University, Kattankulathur 603203, Chennai, Tamil Nadu, India.
Prasanth
P
Bhatt
Institute for Cellular and Molecular Biology, The University of Texas at Austin, 1 University Station A4800, Austin, TX 78712, USA.
Annie J
AJ
Gnanam
Division of Fisheries Biotechnology & Molecular Biology, Department of Biotechnology, Faculty of Science and Humanities, SRM University, Kattankulathur 603203, Chennai, Tamil Nadu, India.
Rajesh
R
Palanisamy
Department of Biotechnology, Faculty of Applied Sciences, AIMST University, Semeling Bedong, 08100 Bedong, Kedah, Malaysia.
Kasi
K
Marimuthu
Lab PCN 206, Microbiology Division, CSIR-Central Drug Research Institute, B.S. 10/1, Sector 10, Jankipuram Extension, Sitapur Road, Lucknow 226031, Uttar Pradesh, India.
Mukesh
M
Pasupuleti
Division of Fisheries Biotechnology & Molecular Biology, Department of Biotechnology, Faculty of Science and Humanities, SRM University, Kattankulathur 603203, Chennai, Tamil Nadu, India. Electronic address: jesuaraj@hotmail.com.
Jesu
J
Arockiaraj
D000371:Agglutination / Q000187:drug effects; D000372:Agglutination Tests; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004912:Erythrocytes / Q000187:drug effects / Q000502:physiology; D005393:Fish Diseases / Q000235:genetics / Q000276:immunology* / Q000382:microbiology; D029941:Fish Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology* / Q000494:pharmacology; D037483:Galectin 1 / Q000737:chemistry / Q000235:genetics* / Q000276:immunology* / Q000494:pharmacology; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D010473:Perciformes / Q000235:genetics* / Q000276:immunology; D010802:Phylogeny; D016415:Sequence Alignment; D014714:Vertebrates / Q000145:classification / Q000235:genetics
D029941:Fish Proteins; D037483:Galectin 1
Bacterial agglutination; Erythrocytes agglutination; Galectin; Gene expression; Murrel