In Silico Prediction and In Vitro Characterization of Unique Function of Human RNase3

Ribonucleases (RNases) constitute a cellular function of exo- and endo-nucleases in all living organisms include vertebrate, bacteria, mold and plant. Human ribonuclease A (hRNaseA) super family consists of thirteen members with high structure similarities. Among which hRNase3 has multiple functions such as RNase, cytotoxicity, and heap ran sulfate (HS) binding activity. Employing Clustal W2 multiple sequence alignment of 13 human RNase A family members revealed that hRNase3 shares 67% identity and 76% similarity with hRNase2, much higher than the others. Besides, both RNase 2 and RNase3 are high conserved among primates. In terms of hRNase3-heparin/heparan sulfate (HS) interaction, three putative heparin binding regions (HBRs), 34RWRCK38 (HBR1), 75RSRFR79 (HBR2) and 101RPGRR105 (HBR3) have been predicted due to presence of positive charge clusters. Furthermore, these regions are also characterized as unique peptide regions within all 13 RNase sequences by REMUS system. Our experimental data derived from protein engineering and binding assays have demonstrated that these HBRs indeed involve in characteristic functions of hRNase3. In summary, through evolution hRNaseA super family has gained novel functions which may be preserved in the unique region or domain to account for additional molecular interactions.