RRC ID 48877
Author Fukuda M, Mikoshiba K.
Title The N-terminal cysteine cluster is essential for membrane targeting of B/K protein.
Journal Biochem J
Abstract B/K protein belongs to a family of C-terminal-type (C-type) tandem C2 proteins that contain two C2 Ca(2+)-binding motifs at the C-terminus. Although other C-type tandem C2 proteins have been found to have a unique N-terminal domain that is involved in membrane anchoring (e.g. synaptotagmin) or specific ligand binding (e.g. rabphilin-3A and Doc2), no research has been conducted on the function of the N-terminal domain of B/K protein. In this study we showed that despite lacking a transmembrane domain, both native and recombinant B/K proteins are tightly bound to the membrane fraction, which was completely resistant to 0.1 M Na(2)CO(3), pH 11, or 1 M NaCl treatment. Deletion and mutation analyses indicated that the cysteine cluster at the N-terminal domain (consisting of seven cysteine residues, Cys-19, Cys-23, Cys-26, Cys-27, Cys-30, Cys-35 and Cys-36) is essential for the membrane localization of B/K protein. When wild-type B/K was expressed in PC12 cells, B/K proteins were localized mainly in the perinuclear region (trans-Golgi network), whereas mutant B/K proteins carrying Cys-to-Ala substitutions were present in the cytosol. Based on our findings, we propose that the N-terminal domain of B/K protein contains a novel cysteine-based protein motif that may allow B/K protein to localize in the trans-Golgi network.
Volume 360(Pt 2)
Pages 441-8
Published 2001-12-1
DOI 10.1042/0264-6021:3600441
PMID 11716773
PMC PMC1222245
MeSH Amino Acid Motifs / genetics Amino Acid Sequence Animals Brain / metabolism COS Cells Calcium-Binding Proteins* Cell Compartmentation / genetics Cysteine / genetics Cysteine / metabolism Cysteine / physiology* Intracellular Membranes / metabolism* Intracellular Membranes / physiology Membrane Glycoproteins / genetics Membrane Glycoproteins / metabolism Membrane Glycoproteins / physiology Mice Molecular Sequence Data Mutagenesis, Site-Directed Nerve Tissue Proteins / genetics Nerve Tissue Proteins / metabolism Nerve Tissue Proteins / physiology* PC12 Cells Protein Structure, Tertiary / genetics Rats Sequence Homology, Amino Acid Synaptotagmins Tandem Repeat Sequences* / genetics
IF 4.097
Times Cited 16
DNA material pEF-T7-mouse SytB/K (RDB15104)