Characterization of human copine III as a phosphoprotein with associated kinase activity.
The copines, first described by Creutz et al. [(1998) J. Biol. Chem. 273, 1393-1402], comprise a two C2 domain-containing protein family and are known to aggregate phosphatidylserine membranes in a calcium-dependent manner. No enzymatic function has been attributed to copines yet. Due to a cross-reacting activity of Mikbeta1, an antibody to the IL-2Rbeta chain, we were able to serendipitously purify, partially microsequence, and clone human copine III. The 5 kb copine III transcript is expressed ubiquitously as determined by a multitissue Northern blot analysis. Phosphoamino acid analysis revealed phosphorylation of copine III on serine and threonine residues. In vitro kinase assays were performed with immunoprecipitated endogenous copine III, chromatography-purified endogenous copine III, and recombinant copine III expressed in Saccharomyces cerevisiae. The exogenous substrate myelin basic protein was phosphorylated in all in vitro kinase assays containing copine III immunoprecipitate or purified copine III. A 60-kDa band was observed in corresponding in gel kinase assays with staurosporine-activated cells. Cell lines expressing high levels of copine III protein had correspondingly high kinase activity in copine III antiserum immunoprecipitate. However, the copine amino acid sequences lack the traditional kinase catalytic domain. Therefore, the data suggest copine III may possess an intrinsic kinase activity and represent a novel unconventional kinase family.
Caudell
E G
EG
Departments of Molecular and Cellular Oncology, Laboratory Medicine, and Head and Neck Surgery, The University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030, USA.
Caudell
J J
JJ
Tang
C H
CH
Yu
T K
TK
Frederick
M J
MJ
Grimm
E A
EA
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004789:Enzyme Activation / Q000235:genetics; D018922:HL-60 Cells; D006801:Humans; D019169:Jurkat Cells; D020014:K562 Cells; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010766:Phosphorylation; D010770:Phosphotransferases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011233:Precipitin Tests; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D020298:U937 Cells
C400933:CPNE7 protein, human; D008565:Membrane Proteins; D010750:Phosphoproteins; C415882:copine III; D010770:Phosphotransferases