Requirement for negative charge on "activation loop" of protein kinase C.
Increasing evidence has implicated a post-translational phosphorylation in the production of a catalytically competent protein kinase C. Here we present structural and biochemical evidence that Thr500 of protein kinase C-beta II is the residue phosphorylated by another kinase. Modeling studies indicate that this residue is part of a "lip" structure at the entrance of the catalytic site; phosphorylation on this lip, or "activation loop," is central to the regulation of three kinases whose structures have been elucidated (Taylor, S. S., and Radzio-Andzelm, E. (1994) Structure 2, 345-355). Biochemical data reveal that mutation of Thr500 to an acidic residue (Glu) results in expression of catalytically active protein kinase C in COS cells. In contrast, mutation of this residue to a neutral, non-phosphorylatable residue (Val) results in expression of inactive enzyme. Thus, negative charge at position 500 is required for catalytically competent protein kinase C-beta II. These data suggest that signal processing by protein kinase C cannot occur until the enzyme is first phosphorylated by a protein kinase C kinase.
Department of Chemistry, Indiana University, Bloomington 47405.
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JW
Orr
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Newton
D000595:Amino Acid Sequence; D001483:Base Sequence; D017931:DNA Primers / Q000737:chemistry; D005971:Glutamates / Q000737:chemistry; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010769:Phosphothreonine / Q000737:chemistry; D011493:Protein Kinase C / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins; D013329:Structure-Activity Relationship; D013912:Threonine / Q000737:chemistry
D017931:DNA Primers; D005971:Glutamates; D011994:Recombinant Proteins; D010769:Phosphothreonine; D013912:Threonine; D011493:Protein Kinase C