Identificação e análise funcional in silico de variantes nos genes de proteínas do leite das raças Guzerá e Gir

Abstract

Milk is a food of high nutritional value and of great importance in human nutrition, a source of vitamins, minerals (especially calcium), carbohydrates, lipids and proteins. Milk consists of two groups of proteins: a solid fraction, formed predominantly by caseins, and a soluble fraction, formed by whey proteins. Caseins are important in cheese production and are a basis for dairy products. Whey proteins bring many benefits to human health, participating in cell repair, generating energy for physical activities and protection for the cardiovascular system. The Brazilian cattle herd is made up of representatives of Taurine (Bos taurus), Zebu (Bos indicus) and their crossbred breeds. Zebu cattle produce most of the national herd with more than 165 million animals, including beef and dairy cattle. Among dairy zebu cattle, the Guzerá and Gir breeds are of great importance for the national agricultural scenario, contributing as producers of milk and meat and as suppliers of genetic material for crossbreeding. The composition of the milk varies according to breed, diet and individual characteristics. Different breeds, throughout their evolution, have accumulated different sets of genetic variants. Some of these variants can be functional, explaining individual or interbreed differences in milk composition. Some functional variants have already been investigated, however, no study has investigated these variants on a genomic scale in the Guzerá and Gir breeds. In this context, the present dissertation had as objectives: 1. To identify, through in silico analyzes, variants in the genes that encode milk proteins, in three Guzerá bulls and three Gir bulls, submitted to whole genome sequencing; 2. Predict, using in silico analysis, the potential for functional repercussion of these variants, that is, to identify which ones should be the target of investigation in future studies, in the search for variants, that would respond for the differences in the concentration of milk proteins between these breeds and between Zebu breeds when compared to taurines. As a result, 70 variants were found in the casein cluster [αS1-casein (CSN1S1), β-casein (CSN2), αS2-casein (CSN1S2) and κ-casein (CSN3)] and in the α-lactalbumin (LALBA) genes, β-lactoglobulin (LGB) and lactoferrin (LTF). Of that total, 37 variants were described for the first time. For functional analysis, 37 new variants and one variant in the coding region already described were selected. Evidence for functionality were found for 20 of these 38 variants: two are missense mutations, two variants affect splicing recognition sites (of these, one also changes a miRNA recognition site), 15 affect an evolutionarily conserved position (of these, two also change miRNA recognition sites) and one altered miRNA recognition site. LTF Thr546Asn and LALBA Ile60Val mutations affect evolutionarily conserved nucleotide positions. Impacts on the 3D structure of the protein due to these mutations were not detected. The mutation in the LTF gene, at position BTA2:53548773, creates an alternative splicing site, which would lead to a premature stop codon, producing an isoform of 550 amino acids. Thus, with this study it was possible to identify 70 variants in the genes that encode milk proteins in the Guzerá and Gir breeds and select 20 potentially functional variants for future studies.