ANTI-SYNAPSIN MONOCLONAL-ANTIBODIES - EPITOPE MAPPING AND INHIBITORY EFFECTS ON PHOSPHORYLATION AND GRB2 BINDING

The synapsins are a family of major neuron-specific synaptic vesicle-a ssociated phosphoproteins which play important roles in synaptic funct ion. In an effort to identify molecular tools which can be used to per turb the activity of the synapsins in in vitro as well as in vivo expe riments, we have localized the epitopes of a panel of monoclonal antib odies (mAbs) raised against synapsins I and II and have characterized their ability to interfere with the interactions of the synapsins with protein kinases, actin and Src homology-3 (SH3) domains. The epitopes of the six mAbs were found to be concentrated in the N-terminal regio n within domains A and B for the synapsin II-reactive mAbs 19.4, 19.11 , 19.51 and 19.21, and in two C-terminal clusters in the proline-rich domains D for synapsin I(mAbs 10.22, 19.51, 19.11 and 19.8) and G for synapsin II (mAb 19.8). The synapsin II-specific mAbs 19.4 and 19.21, whose overlapping epitopes are adjacent to phosphorylation site I, spe cifically inhibited synapsin II phosphorylation by endogenous or exoge nous cAMP-dependent protein kinase. While all the anti-synapsin I mAbs were unable to affect the interactions of synapsin I both with Ca2+/c almodulin-dependent protein kinase II and with actin monomers and fila ments, mAbs 19.8 and 19.51 were found to inhibit the binding of Grb2 S H3 domains to the proline-rich C-terminal region of synapsin I. (C) 19 97 Elsevier Science B.V.