Miller S G, Kennedy M B
J Biol Chem. 1985 Jul 25;260(15):9039-46.
Forebrain and cerebellar Type II Ca2+/calmodulin-dependent protein kinases have different subunit compositions. The forebrain holoenzyme, characterized in our laboratory, is a 650-kDa holoenzyme composed of 50-kDa alpha-subunits and 60-kDa beta-subunits assembled in approximately a 3:1 ratio (Bennett, M. K., Erondu, N. E., and Kennedy, M. B. (1983) J. Biol. Chem. 258, 12735-12744). The cerebellar isozyme is a 500-kDa holoenzyme composed of alpha-subunits and beta-subunits assembled in almost the converse ratio, approximately four beta-subunits for each alpha-subunit. When compared by tryptic peptide mapping and by immunochemical techniques, the beta-subunits from the two brain regions are indistinguishable and the alpha-subunits appear closely related. The specific activities, substrate specificities, and catalytic constants of the cerebellar and forebrain isozymes are similar, suggesting that the alpha- and beta-subunits contain similar catalytic sites. However, two differences in the properties of the isozymes may result in functional differences between them in vivo. First, the apparent affinity of the cerebellar kinase for Ca2+/calmodulin is 2-fold higher than that of the forebrain kinase. Second, the two isozymes appear to associate differently with subcellular structures. Approximately 85% of the cerebellar kinase and 50% of the forebrain kinase remain in the particulate fraction after homogenization under standard conditions. However, they are present in different amounts in postsynaptic density fractions. Postsynaptic densities prepared from forebrain contain the forebrain isozyme. Immunochemical measurements show that it comprises approximately 16% of their total protein. In contrast, postsynaptic densities prepared from cerebellum contain the cerebellar isozyme, but it comprises only approximately 1-2% of their total protein. Thus, the alpha-subunit may play a role in anchoring Type II Ca2+/calmodulin-dependent protein kinase to postsynaptic densities.
前脑和小脑的II型钙/钙调蛋白依赖性蛋白激酶具有不同的亚基组成。我们实验室鉴定的前脑全酶是一种650 kDa的全酶，由50 kDa的α亚基和60 kDa的β亚基以大约3:1的比例组装而成（贝内特，M.K.，埃隆杜，N.E.，和肯尼迪，M.B.（1983年）《生物化学杂志》258，12735 - 12744）。小脑同工酶是一种500 kDa的全酶，由α亚基和β亚基以几乎相反的比例组装而成，即每个α亚基大约有四个β亚基。通过胰蛋白酶肽图谱分析和免疫化学技术比较时，来自两个脑区的β亚基无法区分，α亚基似乎密切相关。小脑和前脑同工酶的比活性、底物特异性和催化常数相似，表明α亚基和β亚基含有相似的催化位点。然而，同工酶性质的两个差异可能导致它们在体内的功能差异。首先，小脑激酶对钙/钙调蛋白的表观亲和力比前脑激酶高2倍。其次，两种同工酶与亚细胞结构的结合方式似乎不同。在标准条件下匀浆后，约85%的小脑激酶和50%的前脑激酶保留在颗粒部分。然而，它们在后突触密度部分的含量不同。从前脑制备的后突触密度含有前脑同工酶。免疫化学测量表明，它约占其总蛋白的16%。相比之下，从小脑制备的后突触密度含有小脑同工酶，但它仅约占其总蛋白的1 - 2%。因此，α亚基可能在将II型钙/钙调蛋白依赖性蛋白激酶锚定到后突触密度中起作用。
