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In Silico Cloning of Green Puffer Fish (tetraodon Nigroviridis) Cysteine Cathepsins and Their Structural Features

M. K. AHMMED
Department of Fishing and Post Harvest Technology, Faculty of Fisheries, Chittagong Veterinary and Animal Sciences University, Chittagong

M. FAISAL
Department of Fishing and Post Harvest Technology, Faculty of Fisheries, Chittagong Veterinary and Animal Sciences University, Chittagong

F. AHMMED
Fisheries and Marine Resource Technology, Khulna University, Khulna

M. R. Ali
Artemia Reference Center, University of Ghent, Belgium

M. N. AHSAN
Fisheries and Marine resource Technology, Khulna University, Khulna, Bangladesh.

Abstract/ Executive Summary:

The study was conducted at the Discipline of Fisheries and Marine Resource Technology, Khulna University, Khulna to know the complete repertoire of Cysteine Cathepsins in Green puffer fish by using Tetraodon genome database through in silico cloning. Cysteine Cathepsins are papain-like Cysteine proteases normally localized in lysosomes. The genomic sequence for a potential Cathepsin gene in Green pufferfish was identified by searching the Tetraodon genome database by using the mature part of some mammalian and fish Cathepsin sequences. The amino acid sequences of some mammalian and fish Cathepsins were collected from the GenBank through National Center for Biotechnology Information (NCBI) website. Stretches of amino acid residues, which are highly conserved among Cathepsins from different species, were used as a query sequence. Complete gene structure predictions have been made by GeneScan. Finally, eight Cysteine Cathepsins were found in Green puffer fish. Each of their eight Tetraodon genes have homologous in other fish mammalian species. Even the relative length of pre, pro and mature domains of Tetraodon Cysteine Cathepsins were found almost equal to representative Cathepsins of other animals reported so far. With the identified Cysteine Cathepsins, multiple sequence comparison and phylogenetic tree construction were carried out and the result revealed that the Tetraodon Cysteine Cathepsins have close evolutionary relationship with the fish Cathepsins, especially with tilapia Cathepsins.

Keywords: Cysteine Cathepsins, Green puffer fish, In silico cloning and GeneScan.

Location: Discipline of Fisheries and Marine Resource Technology, Khulna University, Khulna

Start Date:

End Date:

Program Area: Variety and Species

Commodity: Fish

Objectives:

To know the complete repertoire of Cysteine Cathepsins in Green puffer fish by using Tetraodon genome database through in silico cloning.

Methodology:

The study was conducted at Department of Discipline of Fisheries and Marine Resource Technology, Khulna University, Khulna. The study was based on in silico cloning. Data were collected from secondary sources like different website, journals, Books etc. The genomic sequence of some mammalian and fish Cathepsins, Tetraodon genome database for identifying Tetraodon Cysteine Cathepsins and GenScan for identifying exon intron boundaries and removing introns. ClastalW (http://www.ebi.ac.uk) and GeneDoc software were used for multiple sequence and phylogenetic tree.The genomic sequence for a potential Cathepsin gene in Green puffer fish was identified by searching the Tetraodon genome database using the mature part of known mammalian and fish Cathepsins as the query sequences individually. The amino acid sequences of known mammalian and fish Cathepsins were collected from the GenBank. Then multiple sequences comparison were constructed of similar types of Cysteine Cathepsins of different mammalian and fish species and identified amino acid residues which are highly conserved among Cathepsins from different species. This amino acid residues were also used for query search of Cysteine Cathepsin in Tetraodon genome database. Complete gene structures were predicted through GenScan and exact intron/exon boundaries determined based on GenScan results and fine tune by manual adjustment (http://genes.mit.edu/GENSCAN). Several exons which will be missed by Gene Scan have to be added manually based on the protein DNA alignment. With the identified Cysteine Cathepsins, again multiple sequences comparison was constructed through clastalw website for comparing the sequence structure of different Cathepsins. Phylogenetic tree was developed with guide tree options of clastalw websites to identify evolutionary relationship between the Tetraodon Cysteine Cathepsin and other mammalian and fish Cathepsins.

Source of Information: Int. j. anim. fish. sci. (Online). 3(2): 01-06, April 2015

Article Link (Online Journal):

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Total Budget (Tk.) :

Achievement/Findings:

Eight Cysteine Cathepsins identified in Green puffer fish were found as equal to the other fish and mammalian Cathepsins reported so far. The study also revealed that the Tetraodon Cathepsins have close evolutionary relationship with tilapia rather than other fish species. Thus, comprehensive knowledge about their sequences, structures and functions will help in describing the evolution and biological roles of the entire family. As enzymes are important biological catalyst of various chemical recations and specially Cysteine Cathepsins have application in various pathologies such as cancer, osteoporosis, inflammation and neurological disorders. So, there is a great opportunity of research to identify the cysteine protease family in different organisms.

Knowledge Management: Journal