Inhibition of nerve growth factor‐stimulated neurite outgrowth by methylamine‐modified α₂‐macroglobulin

α₂-Macroglobulin (α₂M) is a rather ubiquitous protein in extracellular spaces of mammals. It is an inhibitor of endopeptidases, can be modified by aliphatic amines, and combines with a number of hormones/cytokines such as β-nerve growth factor (NGF) [Koo PH, Stach RW (1989): J Neurosci Res 22:247]. The objective of this study is to compare the NGF-binding properties of methylamine-modified human α₂M (MA-α₂M) versus normal α₂M and their effects on the biological activity of NGF and neurite extension by embryonic chicken dorsal root ganglia. As determined by gel filtration, polyacrylamide gel electrophoresis, and equilibrium binding studies, these two forms of α₂M are similar in their binding affinities, with MA-α₂M binding about twice as much NGF as normal α₂M. Both normal α₂M and MA-α₂M combine noncovalently with NGF, and prior modification of α₂M is unnecessary for the binding to occur. In contrast to normal α₂M, MA-α₂M potently inhibits the biological activity of NGF and exerts a dose-dependent inhibition on the NGF-stimulated neurite outgrowth by embryonic chicken dorsal root ganglia in culture. The inhibitory effect of MA-α₂M can be overcome by higher NGF concentrations, but is irreversible at lower NGF concentrations. Trypsin- modified α₂M combines covalently and noncovalently with more NGF than normal α₂M but has very little neurite inhibitory activity. The mechanism of inhibition by MA-α₂M is discussed.