XXXV Reunião Anual da SBBqResumoID:8244



The crystallographic structures of the complexes between bovine beta-trypsin and benzamidine derivatives: para-aminobenzamidine and berenil


PERILO, C.S 1; NAGEM, R.A.P 1; PEREIRA, M.T 2; LOPES, J.C.D 3; SANTORO, M.M 1.



1Departamento de Bioquímica e Imunologia - UFMG; 2CDTN - UFMG; 3Departamento de Química - UFMG

Serine proteinases play a central role in several physiologic processes, spanning from digestion to key regulatory mechanisms. Since proteolytic reactions control many biological processes, discovery of new and stronger inhibitors of proteolysis is an important field of modern chemistry. Trypsin is a well-studied serine proteinase specific for carboxyl terminus of lysine and arginine. A large number of synthetic trypsin inhibitors is derived from benzamidine, which is a model system for the basic a -amino acids arginine and lysine. The reactive site of trypsin, as well as its complex with benzamidine has been completely characterized by X-ray crystallography. Thermodynamic studies have shown that p-substituted benzamidine and berenil are even stronger inhibitors of trypsin than benzamidine itself, although the crystallographic structures of these complexes have not been solved yet. In this context, the objective of the present study is to solve the structure of the complexes between beta-trypsin and benzamidine inhibitor derivatives (p-aminobenzamidine and berenil) using X-ray crystallography. Bovine beta-trypsin was purified by chromatography. Crystals of beta-trypsin in complex with p-aminobenzamidine and with berenil were grown at 18 ° C by the hanging drop vapor diffusion method, using ammonium sulfate and/or PEG 3350/4000 as precipitants. Diffraction data were collected at the Instituto de Física de São Carlos (IFSC/USP) using monochromatic radiation from a rotating Cu-anode. Preliminary analyses of electron density maps for both complexes revealed the presence of the ligands in the catalytic cleft of the enzyme. Refinement and structural comparison of both complexes are being conducted at the Instituto de Ciências Biológicas at the Universidade Federal de Minas Gerais (ICB/UFMG).