Angiogenin (ANG), a homologue of bovine pancreatic ribonuclease A (RNase A), promotes angiogenesis (growth of new blood vessels), if present in excess can cause rheumatoid arthritis, psoriasis etc and insufficient amount causes diseases like coronary artery disease, stroke etc. Angiogenin is essential for its angiogenic activity and shows lesser ribonucleolytic activity than RNase A. Therefore, ANG has been suggested as a pharmaceutical target for the development of small-molecule inhibitors of its enzymatic activity as novel antiangiogenic agents. Here, the production of recombinant human ANG in E.coli is achieved using pCMV vector and pET28a vector and purified using Ni-NTA column chromatography. In this study green tea catechins are used as inhibitors of the ribonucleolytic activity of ANG and there affinity and specificity of interaction were also determined using docking studies. The ribonucleolytic inhibitory property of the ligands, Epigallocatechin gallate (EGCG), Epicatechin gallate (ECG) and Gallic acid (GA) were investigated using various spectroscopic techniques. The extent of inhibition was qualitatively confirmed using an agarose gel based assay. From fluorescence and UV-vis spectra, binding constant (Kb) values were calculated which are in the order of 104M-1. Also, the change in β-sheet content of the protein after binding with ligand is revealed from FTIR spectroscopy. Both EGCG and ECG show better ribonucleolytic property compared to GA because of the presence of ring C which links the three aromatic rings. The orientation and conformational flexibility appears to result in a crescent shaped structure that involves possible interactions with the hydroxyl groups on ring B and D with the active site of the protein. The presence of the gallate moiety significantly enhances the ribonucleolytic inhibitory potential that has also been observed in the case of RNase A (previously done in our lab). This structural aspect of polyphenols can be exploited for further studies in the development of anti-angiogenic compounds.

Inhibitory effect of green tea catechins on recombinant human angiogenin

2019 Spring National Meeting

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.