Purification of Different Biologically Active Forms of Mouse β-Nerve Growth Factor by Immobilized Metal Ion Affinity Chromatography

Abstract

This chapter discusses the purification of different biologically active forms of mouse β-nerve growth factor (NGF) by immobilized metal ion affinity chromatography. NGF is a protein that plays a key role in the development and maintenance of neuronal functions in the peripheral and central nervous systems. The biologically active β subunit of the NGF complex is a 26.5 kDa basic protein composed of two identical polypeptide chains linked by strong non-covalent forces. Each chain contains four surface histidines, which shows a high interspecies conservation, making NGF a good candidate for IMAC chromatography. During the purification process from mouse submaxillary gland, NGF is partially modified. The chapter presents an evaluation of the interaction of two different species of NGF with copper, cobalt, nickel, and zinc. This procedure was applied to develop fast purification methods for different N-terminal cleaved forms of mNGF. The role of histidines in the interaction of NGF with metals was also evaluated by diethyl pyrocarbonate modification of protein histidine residues.