Structure of CaMKII

CaMKII was first purified from rat brain homogenates [6,7] and shown by study of its hydrodynamic properties to be a dodecameric hetero-oligomer of two subunits, termed alpha (50 kDa) and beta (60 kDa) [6]. These subunits are highly homologous to each other and are both catalytic.

They appear to assemble together into dodecamers that contain the same average proportions of a- and P-subunits as are present at the time of synthesis. Thus, the holoenzyme composition in a given cell is not homogenous but is instead distributed randomly according to the proportion of available subunits at the time of assembly. There does not appear to be an energetic preference for one holoenzyme composition over another; but this subject has not been studied exhaustively.

Subunits

Cloning of cDNAs encoding subunits of CaMKII demonstrated that the rat genome contains four different genes encoding subunits of CaMKII, each of which has a distinctive pattern of tissue-specific expression (see Table 1). These subunits all apparently associate into dodecameric holoen-zymes, as do the a and P subunits [4]. The most significant differences in sequence among the subunits are found in a region between the amino-terminal catalytic domain (« 300 residues) and the carboxyl-terminal association domain («160 residues) [8]. In this variable region, each of the subunits contains unique sequences ranging from 0 to 70 residues in length; sometimes this region is also alternatively spliced. The variable sequences are believed to confer unique properties. For example, the P-subunit has a higher affinity for calmodulin than does the a-subunit [9,10], perhaps endowing it with greater sensitivity to cytosolic calcium. The P-subunit also displays a much stronger affinity for actin filaments than does the a-subunit [11]. On the other hand, the a-subunit has a higher affinity for the potential postsynaptic density docking protein, densin [12].

Table I Distinct Mammalian Genes Encode Four CaMKII Subunits

Molecular

Subunit weight Tissue distribution

Refs.

54.1 kDa Only in neurons, at very high levels in [8,15] forebrain neurons

60.4 kDa Only in neurons, at moderate levels in [16] most neurons

59 kDa In most tissues, at moderate levels [17,18]

60.1 kDa In most tissues, at moderate levels [17]

Hence, the small differences in properties of the subunits may confer important differences in regulation and subcellular localization.

In the brain, the message encoding the a-subunit is transported into dendrites, whereas that encoding the P-subunit is confined to the soma [13]. Transport of the a-subunit message permits its synthesis in dendrites and, by deduction, assembly of new dendritic "a-only" holoenzymes [14].

Structure of the holoenzyme

Association Domain

Individual subunits associate with each other through their carboxyl-terminal association domains [19]. The two domains mediate formation of an antiparallel dimer. Six dimers then associate to form the dodecameric holoenzyme. The holoenzyme is extremely stable; there is no evidence for the existence of significant amounts of dimers or other intermediate structures in cells, nor is there any indication that holoenzymes can exchange subunits.

Structure Determined by Cryoelectron Microscopy

The individual catalytic domains and the holoenzyme of CaMKII have not been crystallized, but a great deal of insight has come from determination of the structure of a homomeric a-subunit dodecamer at about 3-nm resolution by cryoelectron microscopy [20] (see Fig. 1). This unique structure consists of a hollow, gear-shaped, central cylinder approximately 20 nm in diameter and 10 nm thick. Six slanted flange-like "teeth" project from the surface of the cylinder and confer a six-fold rotational symmetry. Each of the teeth appears to be formed by an antiparallel dimer of the association domains of two subunits. The catalytic domains extend from each end of the teeth to form two parallel rings of six enzymes separated by the cylindrical central structure. The variable regions of the different subunits would form part of the central structure. Thus, specific subcellular association sites located in the variable region would be situated on the central cylinder. The symmetry of the structure suggests that each holoenzyme would contain six pairs of similar binding domains that can mediate subcellular localization. It will be interesting to learn whether this domain arrangement permits the kinase holoenzyme to act as a structural node within the postsynaptic density or other cytoskeletal structures.

Figure 1 Structure of the holoenzyme of CaMKII. The structure of a holoenzyme of a subunits of CaMKII was determined by cryoelectron microscopy by Kolodziej et al. [20]. A surface representation (adapted from Fig. 5 of their paper) is shown in (A). The red central portion is the cylinder formed by the 12 association domains. The yellow, foot-like structures are individual catalytic subunits. Superimposed on two of the catalytic domains is a scaled representation (blue) of the fit of the x-ray structure of the catalytic domain of the cAMP-dependent protein kinase into the surface representation of the "feet." A larger view of this fit (also from Fig. 5 of Kolodjiez et al.) is shown in (B).

Figure 1 Structure of the holoenzyme of CaMKII. The structure of a holoenzyme of a subunits of CaMKII was determined by cryoelectron microscopy by Kolodziej et al. [20]. A surface representation (adapted from Fig. 5 of their paper) is shown in (A). The red central portion is the cylinder formed by the 12 association domains. The yellow, foot-like structures are individual catalytic subunits. Superimposed on two of the catalytic domains is a scaled representation (blue) of the fit of the x-ray structure of the catalytic domain of the cAMP-dependent protein kinase into the surface representation of the "feet." A larger view of this fit (also from Fig. 5 of Kolodjiez et al.) is shown in (B).

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