Binding Efficacy of Heavy Chain Antibody Against Porcine Alpha Amylase Derived from Indian Camel (Camelus dromedarius)

Single domain antibody has huge application in the field of biological sciences. Though many studies have been done on camelid antibody generated from Llama and other camel breeds. In this study single domain antibody is developed from the Indian camel (Camelus dromedarius) in National Research Centre on Camel (NRCC), Jorbeer, Rajasthan, India. The Jaisalmeri breed of camel was immunized with commercially available alpha amylase enzyme. The RNA was isolated from the camel blood and the variable heavy chain antibodies (VHH) region was amplified, cloned in to TG1 strain of E.coli bacteria. Phage display technique was applied to develop peptide library construction. Among different clones, clone A10 was selected on the basis of the binding affinity towards the antigen in vitro. The proteins were further validated by Ramachandran plot using RAMPAGE web server to estimate the degree of nativeness. Best docking results also support the antigen- antibody interaction. This study reveals that dissociation constant, K_D value according to Bobrovnik (2005) equation is 8.22 × 10 ⁻⁹ M. The A10 nanobody is stable even at 95 °C for 30 min. Antigen and antibody interaction is being well characterized by advance bioinformatics tools. These data support the use of therapeutic application in diabetes mellitus to inhibit alpha amylase enzyme. Though there are many drugs available, side effect associated with those drugs can be minimized by the use of our developed anti-alpha amylase single domain antibody. So far, our data has suggested that, in future our developed target antibody sequence can be improved by sequence arrangement towards the cryptic antigen since more target specific nanobody are being discovered.