Journal: The Journal of biological chemistry

Article Title: Distinct Ca2+ and Sr2+ binding properties of synaptotagmins. Definition of candidate Ca2+ sensors for the fast and slow components of neurotransmitter release.

doi: 10.1074/jbc.270.42.24898

Figure Lengend Snippet: FIG. 2. Immobilization of purified full-length synaptotagmin I (lane A) or of the cytoplasmic domains of synaptotagmin I pro- duced by proteolytic cleavage (lane B) on Protein A-Sepharose via a synaptotagmin I-specific monoclonal antibody (Cl41.1). The figure shows a Coomassie Blue-stained SDS gel of the material used for divalent cation-dependent phospholipid binding measurements to brain synaptotagmin I (Fig. 3). Intact synaptotagmin I migrates on the gel as fuzzy monomeric and dimeric bands because it is glycosylated and multimerizes via a sequence close to the transmembrane region (Perin et al., 1991b). Glycosylated sequences and the multimerization domain are not present in the proteolytic cytoplasmic fragment immo- bilized by the antibody (lane B) which consists of the two C2 domains and the C terminus. Filled arrows indicated positions of the different forms of synaptotagmin I. Open arrows point to the heavy and light chain of the monoclonal antibody used for the immobilization. Numbers on the left indicate positions of molecular weight markers.

Article Snippet: Protein concentrations were estimated with a Coomassie Blue-based assay kit (Bio-Rad) or the BCA kit (Pierce).

Techniques: Purification, Staining, SDS-Gel, Binding Assay, Sequencing, Molecular Weight