Marine Antimicrobial Peptides: An Emerging Nightmare to the Life-Threatening Pathogens.

The emergence of multidrug-resistant pathogens due to improper usage of conventional antibiotics has created a global health crisis. Alternatives to antibiotics being an urgent need, the scientific community is forced to search for new antimicrobials. This exploration has led to the discovery of antimicrobial peptides, a group of small peptides occurring in different phyla such as Porifera, Cnidaria, Annelida, Arthropoda, Mollusca, Echinodermata, and Chordata, as a component of their innate immune system. The marine environment, possessing immense diversity of organisms, is undoubtedly one of the richest sources of unique potential antimicrobial peptides. The distinctiveness of marine antimicrobial peptides lies in their broad-spectrum activity, mechanism of action, less cytotoxicity, and high stability, which form the benchmark for developing a potential therapeutic. This review aims to (1) synthesise the available information on the distinctive antimicrobial peptides discovered from marine organisms, particularly over the last decade, and (2) discuss the distinctiveness of marine antimicrobial peptides and their prospects.

In silico molecular and functional characterization of a dual function antimicrobial peptide, hepcidin (GIFT-Hep), isolated from genetically improved farmed tilapia (GIFT, Oreochromis niloticus).

BACKGROUND: Antimicrobial peptides (AMPs), innate immune response molecules in organisms, are also known for their dual functionality, exemplified by hepcidin-an immunomodulator and iron regulator. Identifying and studying various AMPs from fish species can provide valuable insights into the immune profiles of aquaculturally significant fish, which can be made use of in its culture. RESULTS: Hepcidin, a dual-function antimicrobial peptide, was isolated from the gill tissue of Genetically Improved Farmed Tilapia (GIFT-Hep). GIFT-Hep consists of a 90 amino acid pre-propeptide with a 24-mer signal, a 40-mer propeptide, and a 26-mer mature peptide region. The mature peptide had a molecular weight of 3015.61 Da, a theoretical pI of 8.78, a net charge of +4.25, and a protein-binding potential of 2.06 kcal/mol. Four disulfide bonds were formed by eight cysteine residues in the mature region. The presence of positively charged arginine residues renders the peptide 50% hydrophobic. Molecular analysis of GIFT-Hep revealed the presence of a furin propeptide convertase motif, RX(K/R)R, which facilitates trimming of the peptide to yield the mature GIFT-Hep. The hypothetical iron regulatory sequence, QSHLSL, was also identified in the mature peptide. In silico predictions about the characteristics of GIFT-Hep, such as charge, hydrophobicity, high surface accessibility, transmembrane helical regions, hydrophobic faces, hot spots, and cell-penetrating properties, suggest that the peptide functions as an iron regulatory antimicrobial agent. CONCLUSIONS: This study reports a hepcidin antimicrobial peptide with both HAMP1 and HAMP2 properties isolated from genetically improved farmed tilapia, and further evaluation of the properties will prove the feasibility of GIFT-Hep being used as a therapeutant in aquaculture.

A hepatic antimicrobial peptide, hepcidin from Indian major carp, Catla catla: molecular identification and functional characterization.

BACKGROUND: Increase of antibiotic resistance in pathogenic microbes necessitated novel molecules for curing infection. Antimicrobial peptides (AMPs) are the gene-encoded evolutionarily conserved small molecules with therapeutic value. AMPs are considered as an alternative drug for conventional antibiotics. Hepcidin, the cysteine-rich antimicrobial peptide, is an important component in innate immune response. In this study, we identified and characterized hepcidin gene from the fish, Catla catla (Indian major carp) and termed it as Cc-Hep. RESULTS: Open reading frame of Cc-Hep consists of 261 base pair that encodes 87 amino acids. Cc-Hep is synthesized as a prepropeptide consisting of 24 amino acid signal peptide, 36 amino acid propeptide, and 26 amino acid mature peptide. Sequence analysis revealed that Cc-Hep shared sequence similarity with hepcidin from Sorsogona tuberculata. Phylogenetic analysis indicated that Cc-Hep was grouped with HAMP2 family. Structure analysis of mature Cc-Hep identified two antiparallel beta sheets stabilized by four disulphide bonds and a random coil. The mature peptide region of Cc-Hep has a charge of + 2, isoelectric value 8.23 and molecular weight 2.73 kDa. CONCLUSION: Functional characterization predicted antibacterial, antioxidant, and anticancer potential of Cc-Hep, which can be explored in aquaculture or human health care.

Metagenomic analysis of microbial communities in the sediments of a semi-intensive penaeid shrimp culture system.

The present study reports metagenomic sequencing and microbial diversity analysis of the sediment samples of a semi-intensive penaeid shrimp culture system. 16S rRNA gene-based high-throughput sequencing revealed distinct and diverse microbial communities in the analyzed sample. Analysis of the results showed a high abundance of Proteobacteria followed by Verrucomicrobia, Bacteroidetes, Planctomycetes, Firmicutes, Cyanobacteria, and Actinobacteria in the metagenome retrieved from the sediment sample. Unclassified bacteria also contributed a significant portion of the metagenome. Two potential shrimp pathogens viz Vibrio harveyi and Acinetobacter lwoffii detected in the sediment sample show the risk associated with the pond. Microbes that play essential roles in nutrient cycling and mineralization of organic compounds such as Bacteroidetes, Planctomycetes, Gammaproteobacteria, Firmicutes, Cyanobacteria, and Actinobacteria could also be identified. The present study provides preliminary data with respect to the microbial community present in the sediments of a shrimp culture system and emphasizes the application of metagenomics in exploring the microbial diversity of aquaculture systems, which might help in the early detection of pathogens within the system and helps to develop pathogen control strategies in semi-intensive aquaculture systems.

ß-Defensins from common goby (Pomatoschistus microps) and silver trevally (Pseudocaranx georgianus): Molecular characterization and phylogenetic analysis.

Antimicrobial peptides (AMPs) are biologically active molecules involved in host defense present in a variety of organisms. They are an integral component of innate immunity, forming a front line of defense against potential pathogens, including antibiotic-resistant ones. Fishes are proven to be a prospective source of AMPs as they are constantly being challenged by a variety of pathogens and the AMPs are reported to play an inevitable role in fish immunity. Among them, ß-defensins form one of the most studied multifunctional peptides with early evolutionary history and recently being considered as host defense peptides. The present study highlights the first-ever report on ß-defensin AMP sequences from common goby (Pomatoschistus microps) and silver trevally (Pseudocaranx georgianus). A 192 bp cDNA fragment with an open reading frame encoding 63 amino acids (aa) comprising a 20 aa signal peptide region at the N-terminal was obtained from the mRNA of gill tissue of both P. microps and P. georgianus by RT-PCR. These peptide sequences when characterized in silico at the molecular level revealed a 43 aa cationic mature peptide with the signature intra-molecular disulphide bonded cysteine residue pattern ascertaining its ß-defensin identity, further confirmed by phylogenetic analysis. The data collected will pave the way for further research on varied facets of the peptide-like, tissue level expressions, antimicrobial activities on commonly encountered pathogens, and its feasibility as a therapeutant in the aquaculture scenario.

ß-Defensin from the Asian Sea Bass, Lates calcarifer: Molecular Prediction and Phylogenetic Analysis.

Antimicrobial peptides (AMPs) are an important element of the innate immune system of all living organisms and serve as a barrier that safeguards the organisms against a wide range of pathogens. Fishes are proven to be a prospective source of AMPs, and ß-defensins form an important family of AMPs with potent antimicrobial, chemotactic and immunomodulatory activities. The present study reports a ß-defensin AMP sequence (Lc-BD) from the Asian sea bass, Lates calcarifer, a commercially important fish species in tropical and subtropical regions of Asia and the Pacific. A 202-bp cDNA fragment with an open reading frame encoding 63 amino acids (aa) was obtained from the mRNA of gill tissue by RT-PCR. The deduced aa sequence of Lc-BD possessed a signal and a mature peptide region with 20 and 43 aa residues, respectively. Lc-BD was characterized at the molecular level, and a molecular weight of 5.24 kDa and a net charge of +4.5 was predicted for the mature peptide. The molecular characterization of Lc-BD revealed the presence of three intramolecular disulphide bonds involving the six conserved cysteine residues in the sequence, and the phylogenetic analysis of Lc-BD showed a close relationship with ß-defensins from fishes like Siniperca chuatsi, Argyrosomus regius, Trachinotus ovatus and Oplegnathus fasciatus.

A histone H2A derived antimicrobial peptide, Fi-Histin from the Indian White shrimp, Fenneropenaeus indicus: Molecular and functional characterization.

Antimicrobial peptides (AMPs) derived from histone proteins form an important category of peptide antibiotics. Present study deals with the molecular and functional characterization of a 27-amino acid histone H2A derived AMP from the Indian White shrimp, Fenneropenaeus indicus designated as Fi-Histin. This peptide displayed distinctive features of AMPs such as amphiphilic alpha helical structure and a net charge of +6. The synthetic peptide exhibited significant antimicrobial activity against Gram-negative and Gram-positive bacteria especially against V. vulnificus, P. aeruginosa, V. parahaemolyticus, V. cholera and S. aureus. Disruption of cell membrane and cell content leakage were observed in peptide treated V. vulnificus using scanning electron microscopy. The synthetic peptide Fi-His1-21 exhibited DNA binding activity and found to be non-haemolytic at the tested concentrations. Peptide was also found to possess anticancer activity against NCI-H460 and HEp-2 cell lines with an IC50 of 22.670 ±â€¯13.939 µM and 31.274 ±â€¯24.531 µM respectively. This is the first report of a histone H2A derived peptide from F. indicus with a specific antimicrobial activity and anticancer activity, which could be a new candidate for future applications in aquaculture and medicine.

Molecular cloning, recombinant expression and functional characterization of an antimicrobial peptide, Crustin from the Indian white shrimp, Fenneropenaeus indicus.

Antimicrobial peptides (AMPs) comprise molecules that involve in the defense mechanism of various organisms towards pathogens such as bacteria, fungi, parasites and viruses. Crustins are generally defined as multi-domain cationic antimicrobial peptides containing one whey acidic protein (WAP) domain at the C-terminus as the functional unit. In this study, we identified and characterized a novel crustin homolog (Fi-Crustin2) with 354 bp fragment cDNA encoding 117 amino acids and an ORF of 100 amino acids with a net charge of +1 from the mRNA of F. indicus haemocytes. This study forms the second report of a crustin isoform from F. indicus. Blast analysis revealed that Fi-crustin2 exhibits similarity to shrimp crustins already reported. The active mature peptide has a molecular weight of 10.61 kDa and pI of 7.59 with a beta sheeted structure. The mature peptide was cloned into pET-32a(+) with a N-terminal hexa-histidine tag fused in-frame, and expressed in Escherichia coli, and the recombinant crustin, Fi-crustin2 inhibited the growth of Gram-negative bacteria with low MIC. All these features suggest that Fi-crustin2 is a potent antibacterial protein against Gram-negative bacteria and could play an important role in the innate immune mechanism of F. indicus.

Molecular Characterization of a Newly Identified Subfamily Member of Penaeidin from two Penaeid Shrimps, Fenneropenaeus indicus and Metapenaeus monoceros.

Penaeidins are a major group of antimicrobial peptides found in penaeid shrimps. This study reports a new isoform of penaeidin from the hemocytes of Indian white shrimp, Fenneropenaeus indicus (Fi-PEN, JX657680), and the pink shrimp, Metapenaeus monoceros (Mm-PEN, KF275674). Mm-PEN is also the first antimicrobial peptide to be identified from M. monoceros. The complete coding sequences of the newly identified Fi-PEN and Mm-PEN consisted of an ORF of 338 bp encoding 71 amino acids with a predicted molecular weight of 5.66 kDa and a pI of 9.38. The penaeidins had its characteristic signal peptide region (19 amino acids), which was followed by a mature peptide with a proline-rich domain (24 amino acids) at the N-terminal region and a cysteine-rich domain (28 amino acids) at the C-terminal region, designating it to penaeidin-3 subgroup. Structural analysis revealed an alpha-helix in its secondary structure and an extended structure at the proline-rich domain. The newly identified penaeidin isoform showed maximum similarity of 63 % to a penaeidin-3 isoform of P. monodon, which further proves it to be a new isoform. Phylogenetic analysis showed that it possessed similar evolutionary status like other penaeidins, which has subsequently diverged at different phases of evolution. The wide distribution of penaeidins in penaeid shrimps indicates the importance of these AMPs in the innate immunity.

Molecular Characterization and Phylogenetic Analysis of Novel Isoform of Anti-lipopolysaccharide Factor from the Mantis Shrimp, Miyakea nepa.

Anti-lipopolysaccharide factor (ALF) is a cationic anti-microbial peptide representing humoral defence system exhibiting a diverse spectrum of activity against microbial pathogens, including gram-negative and gram-positive bacteria, fungi, parasites and viruses. In this study, we identified and characterized a novel ALF homologue (MnALF) encoding cDNA sequence from the haemocytes of stomatopod mantis shrimp Miyakea nepa. The deduced peptide of MnALF encoded for a 123-amino acid peptide with a 25-residue signal peptide containing selenocysteine followed by a highly cationic mature peptide comprised of a putative LPS-binding domain flanked by two cysteine residues. BLAST analysis of MnALF showed that it exhibits identity to crustacean and limulid ALFs. The mature peptide of MnALF has a net charge of +7 and predicted molecular weight of 10.998 kDa with a theoretical isoelectric point (pI) of 9.93. Spatial structure of MnALF comprises three α-helices packed against a four-stranded ß-sheet of which two were linked by a disulphide bond to form an amphipathic loop similar to the structure of Penaeus monodon, ALF-Pm3. All these features suggest that MnALF could play an imperative role in the innate defence mechanism of M. nepa. To our knowledge, this study accounts for the first report of an anti-microbial peptide from the order stomatopoda.

Immune gene expression profile of Penaeus monodon in response to marine yeast glucan application and white spot syndrome virus challenge.

Immunostimulant potential of eight marine yeast glucans (YG) from Candida parapsilosis R20, Hortaea werneckii R23, Candida spencermartinsiae R28, Candida haemulonii R63, Candida oceani R89, Debaryomyces fabryi R100, Debaryomyces nepalensis R305 and Meyerozyma guilliermondii R340 were tested against WSSV challenge in Penaeus monodon post larvae (PL). Structural characterization of these marine yeast glucans by proton nuclear magnetic resonance (NMR) indicated structures containing (1-6)-branched (1-3)-ß-D-glucan. PL were fed 0.2% glucan incorporated diet once in seven days for a period of 45 days and the animals were challenged with white spot syndrome virus (WSSV). The immunostimulatory activity of yeast glucans were assessed pre- and post-challenge WSSV by analysing the expression profile of six antimicrobial peptide (AMP) genes viz., anti-lipopolysaccharide factor (ALF), crustin-1, crustin-2, crustin-3, penaeidin-3 and penaeidin-5 and 13 immune genes viz., alpha-2-macroglobulin (α-2-M), astakine, caspase, catalase, glutathione peroxidase, glutathione-s-transferase, haemocyanin, peroxinectin, pmCathepsinC, prophenol oxidase (proPO), Rab-7, superoxide dismutase and transglutaminase. Expression of seven WSSV genes viz., DNA polymerase, endonuclease, protein kinase, immediate early gene, latency related gene, thymidine kinase and VP28 were also analysed to detect the presence and intensity of viral infection in the experimental animals post-challenge. The study revealed that yeast glucans (YG) do possess immunostimulatory activity against WSSV and also supported higher survival (40-70 %) post-challenge WSSV. Among the various glucans tested, YG23 showed maximum survival (70.27%), followed by YG20 (66.66%), YG28 (60.97%), YG89 (58.53%), YG100 (54.05%), YG63 (48.64%), YG305 (45.7%) and YG340 (43.24%).

Molecular identification and characterization of Type I crustin isoforms from the hemocytes of portunid crabs, Scylla tranquebarica and Portunus pelagicus.

Crustins are cationic antimicrobial peptides of ca. 7-14kDa with a characteristic four-disulphide core containing WAP domain, present in the hemocytes of crustaceans. The present study reports the first crustin sequences from two portunid crabs, viz. the mud crab Scylla tranquebarica (St-Crustin, JQ965930) and the blue swimmer crab Portunus pelagicus (Pp-Crustin, JQ753312). St-Crustin and Pp-Crustin represented the complete cDNA sequence of Type I crustin, with an ORF of 336bp encoding 111aa with a predicted molecular weight of 10kDa and a pI of 8. The signal sequence contained 21aa residues, which was followed by a mature peptide with a WAP domain at the C-terminus. Peptide model of St-Crustin and Pp-Crustin indicated a randomly coiled structure enclosing two ß-sheets but no helices. St-Crustin and Pp-Crustin shared significant similarities with crustins of portunid crabs (68-95%) and other crabs (60-73%). Phylogenetic analysis showed that St-Crustin and Pp-Crustin possess the same ancestral origin and have a similar evolutionary status like other crustins, which has subsequently diverged at different phases of evolution. St-Crustin and Pp-Crustin were closely related to crab crustins rather than to the crustins of other crustacean groups. The wide distribution of crustins in crustaceans indicates the importance of these AMPs in the innate immunity. Discovery of novel crustins might pave way to the discovery of promising therapeutic/prophylactic agents in health management and disease control in crustacean aquaculture.

Marine yeast Candida aquaetextoris S527 as a potential immunostimulant in black tiger shrimp Penaeus monodon.

A marine yeast Candida aquaetextoris S527 as a source of immunostimulant in Penaeus monodon was studied. Yeast diet was prepared by incorporating 10% C. aquaetextoris S527 biomass into a standard shrimp diet and administered in P. monodon at different frequencies (daily, once in three days, once in seven days and once in ten days) followed by challenge with white spot syndrome virus (WSSV). Immune parameters such as total protein, total hemocyte count, pro-phenoloxidase, nitroblue tetrazolium reduction, alkaline phosphatase activity and acid phosphatase activity were tested. Expression profile of antimicrobial peptide (AMP) genes viz., anti-lipopolysaccharide factor (ALF), crustin-1, crustin-2, crustin-3, penaeidin-3 and penaeidin-5; immune genes viz., alpha-2-macroglobulin (α-2-M), astakine, peroxinectin, prophenol oxidase (proPO) and transglutaminase, and WSSV genes viz., DNA polymerase, endonuclease, protein kinase, immediate early gene, latency related gene, ribonucleotide reductase, thymidine kinase and VP28 were analyzed. The study demonstrated that marine yeast diet administered once every seven days conferred better protection to P. monodon against WSSV infection, supported by the hematological and immune gene expression profiles analyzed.

A Novel Isoform of the Hepatic Antimicrobial Peptide, Hepcidin (Hepc-CB1), from a Deep-Sea Fish, the Spinyjaw Greeneye Chlorophthalmus bicornis (Norman, 1939): Molecular Characterisation and Phylogeny.

Hepcidin is cysteine-rich short peptide of innate immune system of fishes, equipped to perform prevention and proliferation of invading pathogens like bacteria and viruses by limiting iron availability and activating intracellular cascades. Hepcidins are diverse in teleost fishes, due to the varied aquatic environments including exposure to pathogens, oxygenation and iron concentration. In the present study, we report a 87-amino acid (aa) preprohepcidin (Hepc-CB1) with a signal peptide of 24 aa, a prodomain of 39 aa and a bioactive mature peptide of 24 aa from the gill mRNA transcripts of the deep-sea fish spinyjaw greeneye, Chlorophthalmus bicornis. Molecular characterisation and phylogenetic analysis categorised the peptide to HAMP2-like group with a mature peptide of 2.53 kDa; a net positive charge (+3) and capacity to form ß-hairpin-like structure configured by 8 conserved cysteines. The present work provides new insight into the mass gene duplication events and adaptive evolution of hepcidin isoforms with respect to environmental influences and positive Darwinian selection. This work reports a novel hepcidin isoform under the group HAMP2 from a non-acanthopterygian deep-sea fish, C. bicornis.

Two isoforms of anti-lipopolysaccharide factors identified and characterized from the hemocytes of portunid crabs, Portunus pelagicus and Scylla tranquebarica.

Anti-lipopolysaccharide factors (ALFs), a type of cationic antimicrobial peptides (AMPs), and their derivatives are becoming predominant candidates for potential drugs in viral and bacterial diseases. This study reports the first ALF from the mud crab Scylla tranquebarica (StALF, JQ899453) and the second ALF isoform from the blue swimmer crab Portunus pelagicus (PpALF2, JQ899452). Both sequences encoded for precursor molecules, starting with a signal peptide containing 26 amino acid residues, followed by a highly cationic mature peptide, containing two conserved cysteine residues flanking a putative lipopolysaccharide (LPS)-binding domain. BLAST analysis revealed that both PpALF2 and StALF exhibited significant similarity with crustacean ALF sequences. The predicted molecular mass of the mature ALFs was 11.2 kDa with an estimated pI of 10.0. PpALF2 and StALF also showed the typical pattern of alternating hydrophobic and hydrophilic residues in their putative disulphide loop, suggesting that they comprise the same functional domain. Phylogenetic analysis showed that PpALF2 and StALF have similar evolutionary status and they were phylogenetically ancient immune effector molecules which may play an essential role in the host defense mechanism. The spatial structures of PpALF2 and StALF possessed four beta-strands and two alpha-helices. The results indicated that there were more than one ALF involved in crab immunity against various pathogens. ALFs would provide candidate promising therapeutic or prophylactic agents in health management and diseases control in crustacean aquaculture.

Identification of a histone derived, putative antimicrobial peptide Himanturin from round whip ray Himantura pastinacoides and its phylogenetic significance.

Histone H2A participates in host defense responses by producing antimicrobial peptides (AMPs). The present study deals with identification of a putative antimicrobial sequence, Himanturin from the histone H2A of Round Whip Ray, Himantura pastinacoides. A 204 bp fragment encoding 68 amino acid residues was amplified from cDNA of Round Whip Ray, H. pastinacoides. Himanturin exhibited high similarity to previously reported histone H2A derived AMPs indicating the presence of an antimicrobial sequence motif. Physicochemical properties of Himanturin suggest it to be a potential antimicrobial candidate.

Molecular characterization of a crustin-like antimicrobial peptide in the giant tiger shrimp, Penaeus monodon, and its expression profile in response to various immunostimulants and challenge with WSSV.

A crustin-like antimicrobial peptide from the haemocytes of giant tiger shrimp, Penaeus monodon was partially characterized at the molecular level and phylogenetic analysis was performed. The partial coding sequence of 299 bp and 91 deduced amino acid residues possessed conserved cysteine residues characteristic of the shrimp crustins. Phylogenetic tree and sequence comparison clearly confirmed divergence of this crustin-like AMP from other shrimp crustins. The differential expression of the crustin-like AMP in P. monodon in response to the administration of various immunostimulants viz., two marine yeasts (Candida haemulonii S27 and Candida sake S165) and two ß-glucan isolates (extracted from C. haemulonii S27 and C. sake S165) were noted during the study. Responses to the application of two gram-positive probiotic bacteria (Bacillus MCCB101 and Micrococcus MCCB104) were also observed. The immune profile was recorded pre- and post-challenge white spot syndrome virus (WSSV) by semi-quantitative RT-PCR. Expressions of seven WSSV genes were also observed for studying the intensity of viral infection in the experimental animals. The crustin-like AMP was found to be constitutively expressed in the animal and a significant down-regulation could be noted post-challenge WSSV. Remarkable down-regulation of the gene was observed in the immunostimulant fed animals pre-challenge followed by a significant up-regulation post-challenge WSSV. Tissue-wise expression of crustin-like AMP on administration of C. haemulonii and Bacillus showed maximum transcripts in gill and intestine. The marine yeast, C. haemulonii and the probiotic bacteria, Bacillus were found to enhance the production of crustin-like AMP and confer significant protection to P. monodon against WSSV infection.

Molecular characterization and phylogenetic analysis of two antimicrobial peptides: Anti-lipopolysaccharide factor and crustin from the brown mud crab, Scylla serrata.

AMPs are evolutional weapons, widely used by animals and plants in their innate immune system to fend off invading microbes. The present study reports characterization of a new ALF isoform (Sc-ALF; HQ638024) and the first crustin (Sc-crustin; HQ638025) from the mud crab, Scylla serrata. The full-length cDNA of Sc-ALF consisted of 477 bp with an ORF of 123 amino acids and a putative signal peptide of 26 amino acids. Sc-ALF had a predicted molecular weight (MW) of 11.17 kDa and theoretical isoelectric point (pI) of 9.95. Two highly conserved cysteine residues and putative LPS binding domain were observed in Sc-ALF. Comparison of amino acid sequences with neighbor-joining tree indicated that Sc-ALF shared maximum similarity with ALF of S. paramamosain. Peptide model of Sc-ALF created using SWISS-MODEL server was found to consist of two α-helices crowded against a four-strand ß-sheet. The full-length cDNA of Sc-crustin consisted of 433 base pairs with an ORF of 111 amino acids and a putative signal peptide of 21 amino acids. Comparison of amino acid sequences with a neighbor-joining tree revealed that Sc-crustin shared high identity with other known crustins characterized from S. paramamosain, P. trituberculatus, H. araneus, C. maenas and F. chinensis. A whey-acidic-protein domain could be detected at the C-terminus with the characteristic four disulfide core. Sc-crustin had a predicted MW of 10.24 kDa and a pI of 8.76. Peptide model of Sc-crustin created using SWISS-MODEL server indicated a random coiled structure that is with two possible ß-sheets but no helices.