IDENTIFICATION AND CHARACTERIZATION OF DELTA(B)-CAM KINASE AND DELTA(C)-CAM KINASE FROM RAT-HEART, 2 NEW MULTIFUNCTIONAL CA2+/CALMODULIN-DEPENDENT PROTEIN-KINASE ISOFORMS

被引:160
作者
EDMAN, CF [1 ]
SCHULMAN, H [1 ]
机构
[1] STANFORD UNIV, MED CTR, DEPT PHARMACOL, STANFORD, CA 94305 USA
来源
BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH | 1994年 / 1221卷 / 01期
关键词
CALCIUM; KINASE; CALMODULIN; SEQUENCE; EXPRESSION; (RAT HEART);
D O I
10.1016/0167-4889(94)90221-6
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
We have identified, expressed and characterized two new isoforms of the multifunctional Ca2+/calmodulin-dependent kinase (CaM kinase) cloned from rat heart. Both isoforms are variants of the neuronal delta-CaM kinase (termed delta(A)-CaM kinase), and are designated as delta(B)-CaM kinase and delta(C)-CaM kinase. The new isoforms differ from delta(A)-CaM kinase in its isoform-specific insert region, between nucleotides 984 to 1087 of the delta(A)-CaM kinase cDNA. Replacing these 102 nucleotides, a sequence of 33 nucleotides which code for 11 amino acids (KRKSSSSQMM) are introduced in delta(B)-CaM kinase. The delta(C)-CaM kinase lacks all 102 nucleotides and the corresponding 34 amino acids which they encode. The predicted molecular masses of the delta(B)- and delta(C)-CaM kinase isoforms are 57697 Da and 56446 Da, respectively. Recombinant delta-CaM kinases purified from transfected COS-7 cells were found to associate into a larger holoenzyme estimated to contain 8 to 10 subunits. The relative subunit molecular masses on SDS-polyacrylamide gel electrophoresis are 59 kDa, 54 kDa and 52 kDa for delta(A)-, delta(B)- and delta(C)-CaM kinase, respectively. All three isoforms showed a strict dependence on Ca2+/calmodulin for activity and exhibited the Ca2+-dependent autophosphorylation and resultant conversion to Ca2+-independent kinase activity, characteristic features of multifunctional CaM kinase. Phosphopeptide analysis after partial CNBr digestion suggests that delta(B)-CaM kinase is the predominant soluble CaM kinase species purified from rat heart.
引用
收藏
页码:89 / 101
页数:13
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