Analyses of serine proteases and serine protease inhibitors : Evolutionary and structural insights

Analyses of proteases and protease inhibitorsProteolytic enzymes (also known as proteases, proteinases) are enzymes capable of proteolysis (breakdown of proteins), by hydrolysis of the peptide bonds that link amino acids in a polypeptide chain. There are different types of proteases that are found to be important in various biological functions, such as in digestion, immunity, blood clotting etc,. whose deficiency leads to various important diseases such as, hemophilia B, C, neurogenerative disorders etc,.The analyses of proteases in this study consist of three parts, which include1. The sequence and structural analysis of proteases and their inhibitors.2. Development of a residue conservation method to identify members of enzyme families.3. Identification of immunity coding genes and serine protease coding genes in Daphnia magna.Proteolytic enzymes Sequence analysisEvolutionary analysisSerine proteases are proteases which has at least one serine residue in the active site of the enzyme. The aim of this study is to exhibit the ancestral enzyme that had emerged into different classes and subclasses of serine protease.The serpin excluded list of 160 serine proteases of Homo sapiens have been obtained from KEGG database. Each of the selected proteins have been searched for absolute homologues (e-value lesser than or equal to 0.0001) from within all the human proteins using the Blastp program. 97 out of 160 proteins had absolute homologues only from within the serine proteases, which showed that there is high sequence similarity and homology, which suggest that, the classes and subclasses would have emerged from a single ancestral enzyme with a wide-spread cleavage activity. The multiple sequence alignment of the proteins has been performed with clustalW and the phylogenetic analysis with Phylip package. The resulted tree suggested that the different classes and subclasses had a common ancestor, from which they have emerged by different gene duplication events.Future work includes the evolutionary analysis of serine protease inhibitors (serpins) and also finding homologues (of the single ancestral enzyme which would be the result of a sequence clustering technique (described later)) from other organisms and building up the final ancestral enzyme which would have had a wide-spread cleavage activity.Structural analysisNeuroserpinA newly found enzyme, neuroserpin, a selective inhibitor of tPA(tissue-type plasminogen activator) has been taken for the structural an