title: SV31 is a Zn2+-binding synaptic vesicle protein.
abstract: We recently identified in a proteomic screen a novel synaptic vesicle membrane protein of 31 kDa (SV31) of unknown function. According to its membrane topology and its phylogenetic relation SV31 may function as a vesicular transporter. Based on its amino acid sequence similarity to a prokaryotic heavy metal ion transporter we analyzed its metal ion-binding properties and show that recombinant SV31 binds the divalent cations Zn(2+) and Ni(2+) and to a minor extent Cu(2+), but not Fe(2+), Co(2+), Mn(2+), or Ca(2+). Zn(2+)-binding of SV31 in viable cells was verified following heterologous transfection of pheochromocytoma cells 12 (PC12) with recombinant red fluorescent SV31 (SV31-RFP) and the fluorescent zinc indicator FluoZin-3. Sucrose density gradient fractionation of SV31-RFP-transfected PC12 cells revealed a partial overlap of SV31-RFP with synaptic-like vesicle markers and the early endosome marker rab5. Immunocytochemical analysis demonstrated a punctuate distribution in the cell soma and in neuritic processes and in addition in a compartment in vicinity to the plasma membrane that was immunopositive also for synaptosomal-associated protein 25 (SNAP-25) and syntaxin1A. Our data suggest that SV31 represents a novel Zn(2+) -binding protein that in PC12 cells is targeted to endosomes and subpopulations of synaptic-like microvesicles.
authors.affiliation: Institute of Cell Biology and Neuroscience, Biocenter, Goethe-University, Frankfurt am Main, Germany. j.barth@em.uni-frankfurt.de
authors.forename: Joern
authors.initials: J
authors.lastname: Barth
authors.forename: Herbert
authors.initials: H
authors.lastname: Zimmermann
authors.forename: Walter
authors.initials: W
authors.lastname: Volknandt
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D002352:Carrier Proteins / Q000378:metabolism / Q000502:physiology*; D002413:Cations, Divalent / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003198:Computer Simulation; D006224:Cricetinae; D003412:Cricetulus; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D016716:PC12 Cells; D011083:Polycyclic Compounds; D011485:Protein Binding; D011487:Protein Conformation; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D013572:Synaptic Vesicles / Q000378:metabolism / Q000502:physiology*; D014162:Transfection; D015032:Zinc / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D002413:Cations, Divalent; C451182:FluoZin-3; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011083:Polycyclic Compounds; D011994:Recombinant Proteins; C523973:Tmem163 protein, rat; C017800:zinc-binding protein; D015032:Zinc