Association of neurofilament proteins with neuronal Cdk5 activator.
Cdk5 exists in brain extracts in multiple forms, one of which is a macromolecular protein complex comprising Cdk5, neuron-specific Cdk5 activator p35nck5a and other protein components (Lee, K.-Y., Rosales, J. L., Tang, D., and Wang, J.H. (1996) J. Biol. Chem. 271, 1538-1543). The yeast two-hybrid system was employed to identify p35nck5a-interacting proteins from a human brain cDNA library. One of the isolated clones encodes a fragment of glial fibrillary acidic protein, which is a glial-specific protein. Sequence alignment revealed significant homology between the p35nck5a-binding fragment of glial fibrillary acidic protein and corresponding regions in neurofilaments. The association between p35nck5a and neurofilament medium molecular weight subunit (NF-M) was confirmed by both the yeast two-hybrid assay and direct binding of the bacteria-expressed proteins. The p35nck5a binding site on NF-M was mapped to a carboxyl-terminal region of the rod domain, in close proximity to the putative Cdk5 phosphorylation sites in NF-M. A region immediately amino-terminal to the kinase-activating domain in p35nck5a is required for its binding with NF-M. In in vitro binding assays, NF-M binds both monomeric p35nck5a and the Cdk5/p35nck5a complex. The binding of NF-M has no effect on the kinase activity of Cdk5/p35nck5a.
Qi
Z
Z
Department of Medical Biochemistry, University of Calgary, Alberta, Canada.
Tang
D
D
Zhu
X
X
Fujita
D J
DJ
Wang
J H
JH
D000595:Amino Acid Sequence; D000818:Animals; D001923:Brain Chemistry; D002417:Cattle; D051360:Cyclin-Dependent Kinase 5; D018844:Cyclin-Dependent Kinases; D004789:Enzyme Activation; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D016900:Neurofilament Proteins / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D016415:Sequence Alignment
D005904:Glial Fibrillary Acidic Protein; D009419:Nerve Tissue Proteins; D016900:Neurofilament Proteins; C089495:neuronal Cdk5 activator (p25-p35); D051360:Cyclin-Dependent Kinase 5; D017346:Protein Serine-Threonine Kinases; C495905:CDK5 protein, human; D018844:Cyclin-Dependent Kinases