Walter J. Johnson Prize: review
Mechanisms of cyclin-dependent kinase regulation: structures of cdks, their cyclin activators, and cip and INK4 inhibitors1, 2

https://doi.org/10.1006/jmbi.1999.2640Get rights and content

Abstract

The cyclin-dependent kinases (Cdks) have a central role in coordinating the eukaryotic cell division cycle, and also serve to integrate diverse growth-regulatory signals. Cdks are controlled through several different processes involving the binding of activating cyclin subunits, of inhibitory Cip or INK4 subunits, and phosphorylation. Crystallographic studies of Cdks in four different complexes, reviewed here, have revealed the mechanisms by which these regulatory processes control the Cdk switches. All of these mechanisms involve conformational changes in and around the catalytic cleft of the kinase, indicating that Cdks have evolved an intrinsic conformational flexibility. This flexibility is central to their ability to switch states in response to a diverse range of growth-regulatory signals.

Section snippets

The cyclin-dependent kinases

The cell cycle coordinates events needed for the growth of all eukaryotic cells, events such as DNA replication (S phase) and cell division (M phase), ensuring that they occur in the right temporal sequence and proceed in an orderly fashion (reviewed by Sherr 1994, Nurse 1994, King et al 1994). In addition, the cell cycle receives and integrates signals from diverse growth regulatory pathways, ensuring that the cell grows only in the presence of the appropriate signals and in the right

Cdk regulation

Cdks are regulated by several different processes, perhaps reflecting the diversity of the signaling pathways that converge on them. Figure 1(a) summarizes the major regulatory processes common to most Cdks. There are a few additional regulatory processes for subsets of Cdks (reviewed by Morgan, 1995).

When first synthesized, the isolated Cdk subunit has no detectable activity. Its activation occurs in a two-step process. One step is the binding of a cyclin subunit, which imparts partial

The monomeric Cdk

The structure of monomeric Cdk2 has the same overall fold as other eukaryotic protein kinases, first seen in the structure of the cAMP-dependent protein kinase (PKA; Knighton et al., 1991). The structure consists of an N-terminal lobe rich in β-sheet (N lobe), a larger C-terminal lobe rich in α-helix (C lobe), and a deep cleft at the junction of the two lobes that is the site of ATP binding and catalysis (Figure 1(b)). In the monomeric Cdk2 structure, two regions differed from the canonical

Cyclin binding and partial activation

In this step of the activation process, the cyclin binds to one side of the catalytic cleft interacting with both lobes and forming a continuous protein-protein interface (Figure 1(b)). CyclinA contacts to the PSTAIRE helix have a key role in the interface, explaining why cyclin-dependent kinases but not cyclin-independent kinases have this characteristic sequence. Other key contacts are made to the T loop, and parts of the N and C lobes (Jeffrey et al., 1995).

Cyclin binding activates the

Phosphorylation and complete activation

When the T loop becomes phosphorylated on Thr160 (Cdk2), it undergoes an additional conformational change (Figure 3; Russo et al., 1996b). This change is induced by the phosphate group acting as an organizing center in this region, being bound by three arginine side-chains, each coming from a different part of the structure (one from the N lobe, one from the C lobe, and one from the T loop). The arginine residues, in turn, hydrogen bond to other Cdk and cyclin groups, and extend the organizing

Inhibition of the Cdk-cyclin complex by the Cip family

The fully active form of the enzyme can be completely shut down by the binding of the Cip family of inhibitors. One member of the family, p27Cip2, is shown in Figure 1(b) binding the phosphorylated Cdk2-cyclinA complex, interacting with both the Cdk and the cyclin (Russo et al., 1996a).

The obvious mechanism through which p27 inhibits the kinase is through the insertion of a small, 310-helix inside the catalytic cleft (Figure 1(b)). A comparison of the p27-Cdk2-cyclinA and the ATP-Cdk2-cyclinA

Cdk-inhibition by the INK4 family

In the structure of the p16INK4a bound to Cdk6 Russo et al 1998, Brotherton et al 1998, the inhibitor binds next to the catalytic cleft, opposite from where the cyclin would bind, and interacts with both the N and C lobes to form a continuous interface (Figure 1(b)). The INK4 and cyclin binding sites on the Cdk do not overlap, and this explains how INK4 proteins can bind to the Cdk-cyclin complex without dissociating the cyclin.

But if the INK4 and cyclin binding sites do not overlap, how does

Cdks posses an intrinsic conformational flexibility

It is remarkable that all these processes that regulate Cdks at the molecular level work through conformational changes, often so large that they may be better described as structural changes. These recurring structural changes indicate that the Cdk possesses an intrinsic structural flexibility, especially in and around the catalytic cleft (Russo et al., 1998).

Revisiting the mechanisms of Cdk regulation with this intrinsic structural flexibility in mind, it appears that the monomeric Cdk has

Acknowledgements

The author thanks Alicia A. Russo, Philip D. Jeffrey, Lily Tong and Andrea Patten, whose structural studies of the Cdk complexes made this review possible; David O. Morgan for his help with these studies; and Christine Murray for help with the manuscript. The author’s laboratory is supported by the NIH, the Howard Hughes Medical Institute, the Dewitt Wallace Foundation, and the Samuel and Rudin May Foundation.

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