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. 1995 Feb;6(2):161–170. doi: 10.1091/mbc.6.2.161

Molecular cloning and characterization of human kinectin.

A Fütterer 1, G Kruppa 1, B Krämer 1, H Lemke 1, M Krönke 1
PMCID: PMC275826  PMID: 7787243

Abstract

We have identified a human cDNA that is homologous to the chicken kinectin, a putative receptor for the organelle motor kinesin. The human cDNA clone hybridized to a single 4.6-kb mRNA species that codes for a protein of 156 kDa molecular mass. The predicted primary translation product contains an N-terminal transmembrane helix followed by a bipartite nuclear localization sequence and two further C-terminal leucine zipper motifs. In addition, the aminoacid sequence revealed a large region (327-1362) of predicted alpha-helical coiled coils. A monoclonal antibody CT-1 raised against a GST-kinectin fusion protein produced a perinuclear, endoplasmic reticulum-like staining pattern in diverse cell types from different species, indicating evolutionary conservation. Monoclonal antibody CT-1 and anti-chicken kinectin antibodies cross-reacted both in Western blotting and immunoprecipitation with a 160-kDa protein, confirming the antigenic identity of this 160-kDa protein with chicken kinectin. Epitope tagging studies revealed that the nuclear localization sequence motif of kinectin is not functional. Furthermore, a truncated kinesin cDNA lacking the N-terminal hydrophobic domain revealed a nonspecific cytoplasmic staining pattern. Together the data suggest that kinectin is an integral membrane protein anchored in the endoplasmic reticulum via a transmembrane domain.

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