Sequence analysis of UTR and coding region of kappa-casein gene of Indian riverine buffalo (Bubalus bubalis)

Abstract

In this study, complete nucleotide as well as derived amino acid sequence characterization of water buffalo (Bubalus bubalis) kappa-casein gene has been presented. Kappa-casein cDNA clones were identified and isolated from a buffalo lactating mammary gland cDNA library. Sequence analysis of kappa-casein cDNA revealed 850 nucleotides with an open reading frame (ORF) of 573 nucleotides, encoding mature peptide of 169 amino acids. The 5' untranslated region (UTR) comprised 71 nucleotides, while 3' UTR was of 206 nucleotides. A total of 11 nucleotide and seven amino acid changes were observed in, buffalo (Bubalus bubalis) as compared to cattle (Bos taurus), sheep (Ovis aries) and goat (Capra hircus). Among these nucleotide changes, eight were unique in buffalo as they were fully conserved in cattle, sheep and goat. Majority of the nucleotide changes and all the amino acid changes; 14 (Asp-Glu), 19(Asp/Ser-Asn), 96(Ala-Thr), 126(Ala-Val), 128(Ala/Gly-Val), 156(Ala/Pro-Val) and 168(Ala/Glu-Val) were limited to exon IV. Three glycosylation sites, Thr 131, Thr 133 and Thr 142 reported in cattle and goat kappa-casein gene were also conserved in buffalo, however, in sheep Thr 142 was replaced by Ala. Chymosin hydrolysis site, between amino acids Phe 105 and Met 106, important for rennet coagulation process, were found to be conserved across four bovid species. Buffalo kappa-casein with the presence of amino acids Thr 136 and Ala 148 seems to be an intermediate of "A" and "B" variants of cattle. Comparison with other livestock species revealed buffalo kappa-casein sharing maximum nucleotide (95.5%) and amino acid (92.6%) similarity with cattle, whereas with pig it showed least sequence similarity of 76.0% and 53.2%, respectively. Phylogenetic analysis based on both nucleotide and amino acid sequence indicated buffalo kappa-casein grouping with cattle, while sheep and goat forming a separate cluster close to them. The non-ruminant species viz. camel, horse and pig were distantly placed, in separate lineages.

Analysis of genetic variations across regulatory and coding regions of kappa-casein gene of Indian native cattle (Bos indicus) and buffalo (Bubalus bubalis)

“…6 ). A similar finding across different livestock species has been reported by Mukesh et al (2006) based on lineages of nucleotide and amino acid sequences.…”

Identification and Quantification of αS1, αS2, β, and κ-Caseins in Water Buffalo Milk by Reverse Phase-High Performance Liquid Chromatography and Mass Spectrometry

“…The molecular masses of the peaks separated by the described chromatographic protocol were determined by ESI-MS. The comparison between the measured molecular masses and the values obtained from the amino acid sequences of the buffalo proteins ( GenBank accession ) allowed us to clearly identify the peaks corresponding to each casein fraction once the potential posttranslational modifications, in particular phosphorylations, were taken into account. It was possible to identify the peaks shown in Figure as α S1 (peaks 5 and 6), α S2 (peak 2), β (peak 7), and κ-casein (peaks 1, 3, and 4), respectively, based on the results reported in Table .…”

A comprehensive review on the composition and properties of buffalo milk

“…The carbohydrate free fraction of κ-casein represents 40% of κ-CN in buffaloes where it accounted for only 25% of total κ-CN in cow's milk (Addeo et al 1977) which explains the low sialic acid content of buffalo κ-CN. Based on sequence analysis and coding region, Mukesh et al (2006) concluded that the κ-CN of Indian riverine buffalo seems to be an intermediate between A and B variants of cow κ-CN. On the other hand, Abdel Dayem et al (2009) demonstrated that Egyptian buffalo bulls were monomorphic for the κ-CN gene and that they possessed the only B allele.…”