A new derivatizing agent, trimethylammoniopropyl methanethiosulphonate, isefficient for preparation of recombinant brain-derived neurotrophic factorfrom inclusion bodies

Derivatization with trimethylammoniopropyl methanethiosulphonate (TAPS-sulp honate) enabled brain-derived neurotrophic factor (BDNF) to be prepared eff iciently from Escherichia coli inclusion bodies. Reduced BDNF obtained from inclusion bodies solubilized by urea and reduced by dithiothreitol was sug gested to form a complex with itself or with other compounds such as lipids . It could hardly be adsorbed on to cation-exchange resin for partial purif ication prior to a refolding reaction. Reversible derivatization of cystein e residues was tested as a method of dissociating BDNF from such complexes. However, even ifa methyl or aminoethyl group was introduced, BDNF could no t be dissociated readily. Derivatization with TAPS-sulphonate brought about good dissociation of BDNF, and more than 50% adsorbed on to the cation-exc hange resin. BDNF derivatized with TAPS-sulphonate refolded well, and the r efolded samples showed the same biological activity as purified BDNF. Deriv atization with TAPS-sulphonate would increase the intermolecular repulsion of BDNF, due to the positively charged character of the quaternized amine, and inhibit complex formation. Thus, TAPS-sulphonate is effective for the p reparation of BDNF under denatured conditions.