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Cell-free expression and functional reconstitution of CALM in clathrin assembly

Title
Cell-free expression and functional reconstitution of CALM in clathrin assembly
Authors
Kim, JAKim, HL
Ewha Authors
김형래
SCOPUS Author ID
김형래scopus
Issue Date
2001
Journal Title
EXPERIMENTAL AND MOLECULAR MEDICINE
ISSN
1226-3613JCR Link
Citation
vol. 33, no. 2, pp. 89 - 94
Keywords
expressionclathrin-coated vesicleCALMSH3 domaincleavage
Publisher
KOREAN SOC MED BIOCHEMISTRY MOLECULAR BIOLOGY
Indexed
SCI; SCIE; SCOPUS; KCI WOS scopus
Abstract
Clathrin-mediated vesicle formation is an essential step in the intracellular trafficking of the protein and lipid. Binding of clathrin assembly protein to clathrin triskelia induces their assembly into clathrin-coated vesicles (CCVs). In order to better understand a possible role of post-translational modification of CALM (clathrin assembly protein lymphoid myeloid), the homologue of AP180, in the assembly of CCVs, CALM was expressed in the cell-free reticulocyte translation system that is capable of carrying out post-translational modification. The apparent molecular weight of the expressed recombinant CALM was estimated as 105 kD. Alkaline phosphatase treatment of CALM resulted in a mobility shift on SDS-PAGE. We found that CALM was associated with the proteins harboring SH3 domain, promote assembly of clathrin triskelia into clathrin cage and bound to the preformed clathrin cage. CALM was also proteolyzed by caspase 3 and calpain but not by caspase 8. These results indicated that the post-translationally modified CALM, expressed in the eukaryotic cell-free reticulocyte translation system was able to mediate the assembly of clathrin and the coated-vesicle formation.
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의학전문대학원 > 의학과 > Journal papers
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