Urea-Induced Dissociation of Nerve Growth Factor from Venom of Chinese Cobra by SEC

摘要

The urea-induced dissociation of nerve growth factor from venom of Chinese cobra (_cNGF) was studied by intrinsic fluorescence emission spectra, SEC, urea-gradient polyacrylamide gel electrophoresis, assays of biological activity and thermodynamic parameters. The results showed that when urea concentration was lower than or equal to 4.0 mol L~(-1) or higher than or equal to 8.0 mol L~(-1), _cNGF existed only in native homodimer form or monomer form, respectively; whereas when urea concentration was higher than 4.0 mol L~(-1) and lower than 8.0 mol L~(-1), they existed simultaneously in the native homodimer and monomer forms and the former decreased, while the latter increased with the increase in urea concentration. Based on the association-dissociation equilibrium between _cNGF and urea molecules, an equation, which includes two characteristic dissociation parameters K and Δm, was presented to describe the urea-induced dissociation process of _cNGF. As the reaction temperature increased from 15 to 35 °C, positive enthalpy and entropy changes were observed, and the parameter K increased from 2.72×10~(-13) to 5.18×10~(-12) (L?mol~(-1)), while the parameters Δm and ΔG, respectively, decreased from 10.18 to 8.42 and from -10.27 to -18.67 (kJ mol~(-1)), which means that the urea-induced dissociation of _cNGF was spontaneous and entropy-driven and the higher temperature was favorable for the dissociation process. Using the procedures and equations mentioned in the paper, the urea-induced dissociation of _cNGF is first comprehensively described. Furthermore, this work presents a useful method for people to study the dissociation of dimer or multimer proteins induced by denaturants, inducers, pH, etc.