BIOINFORMATIC ANALYSES OF KAPPA CASEIN (CSN3) GENE IN TWENTY-FOUR MAMMALIAN SPECIES

ABSTRACT Kappa casein (CSN3) gene is a variant of the milk protein highly conserved in mammalian species. Genetic variations in CSN3 gene of 24 mammalian species were investigated using bioinformatic approach. A total of 55 CSN3 gene sequences with corresponding amino acids belonging to the 24 species were retrieved from GenBank. Using a comparative genomic approach, 1997 base pairs (bp) of the CSN3 sequences belonging to 14 homologous species and 10 divergent taxa were obtained. Alignment of 55 sequences within the region of 1498 bp and containing 499 gaps were carried out using Clustal W which revealed high polymorphisms of CSN3 sequences among the species. The percent identity and similarity between species were determined by conducting pair-wise comparisons of the sequences which revealed high level of identity and similarity among the ruminants (81-99% and 82-100%) except for camel. This implied that the gene is conserved in all the ruminants. However, percent identity and similarity (33-99% and 53-99%) between mammalian non-ruminant species were lower than values from mammalian ruminant species. This revealed that CSN3 protein from them might be functionally different and that divergence among them is ancient. Higher polymorphisms of amino acids were observed among the species than within species. Four non-synonymous mutations occurred in cattle, six in goats, four in horses, three in rabbits and seven in sheep. The results of DnaSP analysis indicated that the selected regions (1 to 1997 bp) of the fifty-five sequences from all the species had 1498 nucleotide sites excluding sites with gaps (499). There are invariable and variable sites that include 185 singleton variable sites and 56 parsimony informative sites. The nucleotide diversity (π = 0.212) and average number of nucleotide diversity (k = 94.84) for all sequences which were lower than the highest values in rabbit (π = 0.557, k = 229) were estimated. Divergence of the species could be inferred from high genetic diversity of the CSN3 gene. The haplotype diversity was 1.00 in mouse, rabbits, goats, horses, camel, sheep and human indicating abundant genetic diversity in those species. Close relationship among members of the same genus (Bos and Equid) was observed in constructed phylogenetic trees of CSN3 gene which showed that the comparability of CSN3 gene sequences was highest among these members and they evolved from a most recent common ancestor. Comparison of the prediction of secondary structures of CSN3 using different six bioinformatic software revealed that the best prediction is from GOR IV and PREDATOR, respectively. Both GOR IV and PREDATOR revealed that the primary sequence of the protein basically has three segments: helix, strand and coil. The tertiary structure produced from the PHYRE software appeared to be the most likely structure from protein homology to the database. SWISS MODEL software could not predict the tertiary structure due to inability of the tool to find appropriate protein of known structure as template for the modelling. A more detailed study involving complete coding sequences of the gene and wet laboratory work will help to delineate how the mutations affect the functions of the gene.