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Don Anushka Sandaruwan Elvitigala,Jehee Lee 제주대학교 해양과학연구소 2021 해양과환경연구소 연구논문집 Vol.45 No.-
Cystatins are reversible inhibitors of cysteine proteases which show an omnipresent distribution in the life on earth. Although, cystatins with mammalian origin were well characterized and their roles in physiology were reported in details, those from teleostean origin are still underrepresented in literature. However, role of cystatins in fish physiology and immune defense is highlighted in few recent reports. In this study, a cystatin C holmologue from rock bream (Oplegnathus fasciatus); termed RbCytC was identified and molecularly characterized. The complete coding sequence of RbCytC was 387 bp in length, which codes for a polypeptide with 129 amino acids, including a signal peptide of 19 amino acids. The consensus cystatin family signatures including a G residue, turning up towards the N-terminus region, QVVAG motif, locating at the middle of the sequence and the PW motif at the c terminal region was found to be well conserved in RbCytC. Phylogenetic analysis using different cystatin counterparts affirmed the close evolutionary relationship of RbCytC with its teleostan homologs which belong to family 2 cystatins. The predicted molecular model of RbCytC resembled most of the structural features of empirically elucidated tertiary structures for chicken egg white cystatin. According to the qPCR assays, RbCytC showed detectable expression in all fish tissues used in the experiment, with markedly pronounced expression level in liver. Moreover, its basal mRNA expression was up-regulated in liver and spleen tissues by experimental rock bream iridovirus infection, whereas down regulated in the same tissues, post live Edwardsiella tarda injection. Collectively, outcomes of our study validate the structural homology of RbCytC with known cystatin C similitudes, especially those of teleosts and suggest its potential roles in proteolytic processes of rock bream physiology as well as in host immune defense mechanisms.
Elvitigala, Don Anushka Sandaruwan,Priyathilaka, Thanthrige Thiunuwan,Whang, Ilson,Nam, Bo-Hye,Lee, Jehee Elsevier 2015 FISH AND SHELLFISH IMMUNOLOGY Vol.44 No.1
<P><B>Abstract</B></P> <P>Antioxidative defense renders a significant protection against environmental stress in organisms and maintains the correct redox balance in cells, thereby supporting proper immune function. Catalase is an indispensable antioxidant in organisms that detoxifies hydrogen peroxides produced in cellular environments. In this study, we sought to molecularly characterize a homolog of catalase (RfCat), identified from black rockfish (<I>Sebastes schlegelii</I>). <I>RfCat</I> consists of a 1581?bp coding region for a protein of 527 amino acids, with a predicted molecular weight of 60?kD. The protein sequence of RfCat harbored similar domain architecture to known catalases, containing a proximal active site signature and proximal heme ligand signature, and further sharing prominent homology with its teleostan counterparts. As affirmed by multiple sequence alignments, most of the functionally important residues were well conserved in RfCat. Furthermore, our phylogenetic analysis indicates its common vertebrate ancestral origin and a close evolutionary relationship with teleostan catalases. Recombinantly expressed RfCat demonstrated prominent peroxidase activity that varied with different substrate and protein concentrations, and protected against DNA damage. <I>RfCat</I> mRNA was ubiquitously expressed among different tissues examined, as detected by qPCR. In addition, <I>RfCat</I> mRNA expression was modulated in response to pathogenic stress elicited by <I>Streptococcus iniae</I> and poly I:C in blood and spleen tissues. Collectively, our findings indicate that RfCat may play an indispensable role in host response to oxidative stress and maintain a correct redox balance after a pathogen invasion.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Homolog of catalase was identified from black rockfish (RfCat) RfCat. </LI> <LI> RfCat resembled typical catalase domain architecture. </LI> <LI> Recombinant RfCat (rRfCat) showed detectable peroxidase activity. </LI> <LI> rRfCat could notably protect bacterial cells and DNA from oxidative damage. </LI> <LI> Transcriptional level of RfCat was modulated under pathogenic stress. </LI> </UL> </P>
Elvitigala, Don Anushka Sandaruwan,Thulasitha, William Shanthakumar,Lee, Jehee Springer-Verlag 2016 Genes & Genomics Vol.38 No.3
<P>Nucleotide-oligomerization domain like receptors (NLRs) are recently identified group of pattern recognition receptors which involve in sensing broad range of pathogen associated molecular patterns or damage associated molecular patterns to trigger corresponding immune responses in host cells. In this study, we identified and characterized a NLRC5 family member from a previously established black rockfish cDNA database, designating as 'RfNLRC5'. The complete open reading frame of RfNLRC5 consists of 5808 bp which encodes for a protein of 1936 amino acids with the predicted molecular mass of 213 kDa. Intriguingly, RfNLRC5 harbored only two typical domain signatures of NLR superfamily, namely NACHT domain and LRRs. However, it was phylogenetically closely related to the telostan counterparts. As expected, RfNLRC5 shared significant sequence compatibility with its teleostan counterparts, eminently with that of large yellow croaker. As detected by our qPCR assay, RfNLRC5 was universally distributed in tissues examined, albeit with different levels. Therein, more pronounced expression levels were detected in blood cells and spleen tissues. After treating the na < ve fish with immune stimulants; lipopolysaccharides and Polyinosinic:polycytidylic acid (poly I:C), RfNLRC5 mRNA expression in blood cells and spleen tissues was found to modulate significantly with notable inductive responses. Collectively, our results in this study hint a potential role of RfNLRC5 in host innate immune responses against bacterial or viral infections.</P>
Jo, Eunyoung,Elvitigala, Don Anushka Sandaruwan,Wan, Qiang,oh, Minyoung,Oh, Chulhong,Lee, Jehee Elsevier 2017 DEVELOPMENTAL AND COMPARATIVE IMMUNOLOGY Vol.77 No.-
<P><B>Abstract</B></P> <P>Dendritic-cell-specific ICAM-3-grabbing non-integrin (DC-SIGN) is a C-type lectin that functions as a pattern recognition receptor by recognizing pathogen-associated molecular patterns (PAMPs). It is also involved in various events of the dendritic cell (DC) life cycle, such as DC migration, antigen capture and presentation, and T cell priming. In this study, a DC-SIGN-like gene from the big belly seahorse <I>Hippocampus abdominalis</I> (designated as <I>ShDCS-like</I>) was identified and molecularly characterized. The putative, complete ORF was found to be 1368 bp in length, encoding a protein of 462 amino acids with a molecular mass of 52.6 kDa and a theoretical isoelectric point of 8.26. The deduced amino acid sequence contains a single carbohydrate recognition domain (CRD), in which six conserved cysteine residues and two Ca<SUP>2+</SUP>-binding site motifs (QPN, WND) were identified. Based on pairwise sequence analysis, ShDCS-like exhibits the highest amino acid identity (94.6%) and similarity (97.4%) with DC-SIGN-like counterpart from tiger tail seahorse <I>Hippocampus comes.</I> Quantitative real-time PCR revealed that <I>ShDCS-like</I> mRNA is transcribed universally in all tissues examined, but with abundance in kidney and gill tissues. The basal mRNA expression of <I>ShDCS-like</I> was modulated in blood cell, kidney, gill and liver tissues in response to the stimulation of healthy fish with lipopolysaccharides (LPS), <I>Edwardsiella tarda,</I> or <I>Streptococcus iniae</I>. Moreover, recombinant ShDCS-like-CRD domain exhibited detectable agglutination activity against different bacteria. Collectively, these results suggest that ShDCS-like may potentially involve in immune function in big belly seahorses.</P> <P><B>Highlights</B></P> <P> <UL> <LI> DC-SIGN as a pattern recognition receptor by recognizing pathogen-associated molecular patterns (PAMPs). </LI> <LI> DC-SIGN-like was identified from big belly seahorse (<I>Hippocampus abdominalis</I>) (ShDCS-like). </LI> <LI> ShDCS-like has characteristic of C-type lectin. </LI> <LI> ShDCS-like ubiquitously expressed in wide range of tissues and stimulated under pathogenic stress signals. </LI> </UL> </P>
Lee, Seongdo,Elvitigala, Don Anushka Sandaruwan,Lee, Sukkyoung,Kim, Hyun Chul,Park, Hae-Chul,Lee, Jehee Elsevier 2017 Developmental and comparative immunology Vol.67 No.-
<P><B>Abstract</B></P> <P>Bactericidal permeability-increasing protein (BPI)/lipopolysaccharide (LPS) binding proteins (LBPs) are well-known proteins that play an indispensable role in host antimicrobial defense. Herein, we report a homolog of BPI/LBP from black rockfish (<I>Sebastes schlegelii</I>) (designated as RfBPI/LBP) and characterize its structural and functional features at the molecular level. We identified the putative complete open reading frame (1422 bp) of <I>RfLBP</I> that encodes a 474 amino acid protein with a predicted molecular mass of ∼51.5 kDa. The primary protein sequence of RfBPI/LBP contains domain features of BPI/LBP family proteins and shares significant sequence consistency with its homologs. Our phylogenetic analysis clearly demonstrated the vertebrate ancestral origin of RfBPI/LBP, further reinforcing its evolutionary relationship with teleostean homologs. Recombinant RfBPI/LBP demonstrated <I>in vitro</I> LPS-binding activity and antibacterial activity against <I>Escherichia coli</I>, but not against <I>Streptococcus iniae</I>. Moreover, <I>RfBPI/LBP</I> exhibited temporal transcriptional activation against pathogens and pathogen-associated molecular patterns. Collectively, our findings suggest that RfBPI/LBP plays an essential role in host antimicrobial defense, plausibly through selective eradication of invading bacteria.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Homolog of BPI/LBP was identified from black rockfish (RfBPI/LBP). </LI> <LI> Rf RfBPI/LBP resembled typical domain architecture of its homologues. </LI> <LI> Recombinant RfBPI/LBP showed selective antibacterial and LPS binding activity. </LI> <LI> <I>RfBPI/LBP</I> was ubiquitously expressed in tissues under physiological conditions. </LI> <LI> Transcriptional level of <I>RfBPI/LBP</I> was modulated under pathogenic stress. </LI> </UL> </P>
이영득,이지현,심원보,Don Anushka Sandaruwan Elvitigala,디조이사마하나마,이수진,허수진,이제희,강도형,오철홍 한국해양과학기술원 2014 Ocean science journal Vol.49 No.4
Glucanases are involved in degradation of glucans. Here, we report a new endo-β-1,3-glucanase Mzl86 identified in Mesoflavibacter zeaxanthinifaciens S86. The deduced amino-acid sequence of Mzl86 showed highest similarity (45.1%) with Leeuwenhoekiella blandensi and thus placed in glycosyl hydrolase family 16. Purified recombinant protein (rMz186) showed an optimum enzyme activity against laminarin at 50°C and pH 8. The enzyme was stable at 50°C for 1 hour (maintaining 80% of its maximum activity) and was strongly activated (187%) in the presence of 2.5 mM manganese. Substrate-specific activities of rMzl86 against laminarin, barley β-glucan and lichenan were 261, 128 and 115 unit/mg, respectively. rMzl86 degraded laminarioligosaccharides (lager than biose) and laminarin while producing mainly biose and glucose. Molecular and biochemical properties reveal that rMzl86 shares typical features of β-1,3-glucanase (EC 3.2.1.39) and thus is a potential candidate for use in agriculture, drug, chemical and bioethanol industries.
Future Prospects and Health Benefits of Functional Ingredients from Marine Bio-resources: A review
Samarakoon, Kalpa W.,Elvitigala, Don Anushka Sandaruwan,Lakmal, H.H. Chaminda,Kim, Young-Mog,Jeon, You-Jin The Korean Society of Fisheries and Aquatic Scienc 2014 Fisheries and Aquatic Sciences Vol.17 No.3
The marine ecosystem represents a vast and dynamic array of bio-resources attributed with its huge diversity and considered as potential untapped reservoirs for the development of functional foods for future health markets. Basically, marine microorganisms, sponges, algae, invertebrates such as crustaceans and mollusks along with marine fish species can be considered as marine bio-resources, which can be utilized to obtain different health benefits for humans, directly or after processing. Most of the bio-molecular components, such as lipids and proteins from these marine bio-resources, which can be extracted in large scale using the modern and advanced biotechnological approaches, are suitable drug candidates for the pharmaceutical industry as well as functional food ingredients for the food industry. Moreover, the furtherance of high throughput molecular biological techniques has already been incorporated with identification, mining and extraction of molecular components from marine bio-resources. In this review, potential marine bio-resources with respect to their extractable bio-molecules were described in details, while explaining the present and prospective methods of identification and extraction, which are integrated with advanced techniques in modern biotechnology. In addition, this provides an overview of future trends in marine biotechnology.