The N-terminal cysteine cluster is essential for membrane targeting of B/K protein

M. Fukuda, K. Mikoshiba

Research output: Contribution to journalArticlepeer-review

17 Citations (Scopus)

Abstract

B/K protein belongs to a family of C-terminal-type (C-type) tandem C2 proteins that contain two C2 Ca2+-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 Na2CO3, pH 11, or 1 M NaC1 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.

Original languageEnglish
Pages (from-to)441-448
Number of pages8
JournalBiochemical Journal
Volume360
Issue number2
DOIs
Publication statusPublished - 2001 Dec 1
Externally publishedYes

Keywords

  • C-type tandem C2 protein
  • C2 domain
  • Cys cluster
  • Synaptotagmin

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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