AFFINITY PURIFICATION AND SUBCELLULAR-LOCALIZATION OF KINESIN IN HUMAN NEUTROPHILS

Citation
Sw. Rothwell et al., AFFINITY PURIFICATION AND SUBCELLULAR-LOCALIZATION OF KINESIN IN HUMAN NEUTROPHILS, Journal of leukocyte biology, 53(4), 1993, pp. 372-380
Citations number
24
Categorie Soggetti
Immunology,Hematology
ISSN journal
07415400
Volume
53
Issue
4
Year of publication
1993
Pages
372 - 380
Database
ISI
SICI code
0741-5400(1993)53:4<372:APASOK>2.0.ZU;2-V
Abstract
Studies of granule-microtubule interactions in human neutrophils have suggested that mechanochemical ATPases such as kinesin or dynein may p lay a role in granule mobilization during neutrophil activation by inf lammatory signals. In this study we show that proteins extracted from the surface of neutrophil granules, found previously to contain microt ubule-dependent ATPase activity, caused microtubules polymerized from phosphocellulose-purified rat brain tubulin to move across glass slide s. Antibodies were generated against peptides based on two regions of the amino acid sequence of Drosophila kinesin: the ATPase active site (amino acids 86-99) in the head of the kinesin heavy chain and the tai l of the heavy chain (residues 913-933). These antibodies were found t o recognize kinesin in rat brain extracts as well as kinesin-like poly peptides in extracts of human neutrophils. Furthermore, when used in i mmunoaffinity chromatography, these antibodies permitted the isolation of a protein from neutrophil granule extracts that was recognized by Drosophila kinesin antibodies. Subcellular localization by immunofluor escence microscopy showed this protein to be associated principally wi th the cytoplasmic granules of neutrophils.