Fig. 7. The C2B of synaptotagmin VII contains a dominant and transplantable inhibitory subdomain. Chimeric constructs were generated that replaced ß-strands of the eight-stranded ß-barrel in pairs. The resulting chimeras fell into two categories: a synaptotagmin VII C2B with (A) increasing numbers of N-terminal synaptotagmin-I-derived ß-strands [7B series (F7B-H7B)] and (B) the inverse series [1B series (F1B-H1B)]. The construct compositions are as follows: F7B (synaptotagmin I residues 266-303 fused to synaptotagmin VII residues 298-403), G7B (synaptotagmin I residues 266-334 fused to synaptotagmin VII residues 329-403), H7B (synaptotagmin I residues 266-369 fused to synaptotagmin VII residues 364-403), F1B (synaptotagmin VII residues 261-297 fused to synaptotagmin I residues 304-421), G1B (synaptotagmin VII residues 261-328 fused to synaptotagmin I residues 335-421), H1B (synaptotagmin VII residues 261-363 fused to synaptotagmin I residues 370-421). PC12 cells expressing these chimeras were subsequently studied for their capacity to endocytose the proteins with the endocytosis assay. (C) A model of the synaptotagmin VII C2B domain depicts the relative positions of the inhibitory region (red), the AP-2-binding site (cyan) and the WHXL motif (green).

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