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Cdk5/p25 nck5a interaction with synaptic proteins in bovine brain
Author(s) -
Rosales Jesusa L.,
Nodwell Michael J.,
Johnston Randal N.,
Lee KiYoung
Publication year - 2000
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/(sici)1097-4644(20000701)78:1<151::aid-jcb14>3.0.co;2-l
Subject(s) - cyclin dependent kinase 5 , amphiphysin , immunoprecipitation , size exclusion chromatography , chemistry , microbiology and biotechnology , biochemistry , kinase , biology , enzyme , protein kinase a , endocytosis , receptor , cyclin dependent kinase 2 , dynamin , gene
Cyclin‐dependent kinase 5 (Cdk5) exists in large multimeric complexes, but its function and binding partners in these complexes are unclear. We explored these issues by chromatographic and immunochemical analyses of Cdk5 and p25 nck 5 a (a neuronal Cdk5 activator) and their associated proteins from bovine brain. Mono‐S column enzyme eluates were divided into three fractions and analyzed by gel filtration. The majority of p25 nck 5 a from Mono‐S fractions I, II, and III eluted from the gel filtration column at ∼60, 200, and 400 kDa, respectively, and Cdk5 was abundant in fractions >400 kDa. We characterized these macromolecular structures by immunoprecipitating p25 nck 5 a , followed by a second immunoprecipitation of remaining unbound proteins using a Cdk5 antibody. The p25 nck 5 a immunoprecipitates showed association with Cdk5. Amphiphysin was detected in the 400‐kDa complex and synapsin I in the >400 kDa structure. The Cdk5 immunoprecipitates, however, revealed abundant retained Cdk5 but no remaining p25 nck 5 a , indicating that Cdk5 in macromolecular structures is mostly unassociated with p25 nck 5 a . Thus, we demonstrate: an amphiphysin‐associated 400‐kDa Cdk5/p25 nck 5 a complex, a synapsin I‐associated >400‐kDa Cdk5/p25 nck 5 a complex, and nck5a‐free Cdk5 complexes (200 to >400 kDa). Amphiphysin acts as a Cdk5/p25 nck 5 a substrate in the 400‐kDa complex and we speculate that Cdk5/p25 nck 5 a participates in amphiphysin‐mediated endocytosis. J. Cell. Biochem. 78:151–159, 2000. © 2000 Wiley‐Liss, Inc.