Kinases phosphate binding region

An important observation by Naumann and Matter is the identification of additional opportunities to achieve selective interactions in the kinases phosphate binding region, whereas most work so far has focussed on the purine binding regions. 

At present there are >100 protein kinase-inhibitor structures in publidy available structural databases which span 28 kinases and a variety of inhibitor structural classes. Based upon an analysis of these data a classification of ATP binding regions has been proposed by Traxler and Furet [59] and Bower (personal communication from Michael J. Bower, December 1999). In the subsequent discussion a slightly modified version of this classification will be used to organize the trends seen across kinases and inhibitor dasses (see Fig. 2.2). In the Traxler model, five sites were proposed of which three (adenine, sugar and phosphate-binding sites) can be directly related to ATP and two additional lipophilic sites which lay outside of the region occupied by ATP. In the Bower model, an additional polar site on the surface of the protein was proposed. 

Interpretation of the structural features responsible for this landscape reveals that the main differences along PC 1 (i.e. between PKA and MAP/CDK kinases) are found in the purine and hinge binding region, whereas the discrimination in PC 2 is mainly driven by structural differences in the phosphate binding area. 

The ATP-binding region of the receptor is located around Lys-1018 (Ebina etal., 1987) and Gly-996 (Odawara etal., 1989). The cytoplasmic sequence of the insulin receptor contains 13 tyrosine residues and it is believed that at least six of these tyrosines become phosphorylated (White et al., 1984, 1985c, 1988a Tornqvist et al., 1987, 1988 Tavare and Denton, 1988 Tavare et al., 1988 Tornqvist and Avruch, 1988) after insulin stimulation. The triplet of tyrosines at 1146, 1150 and 1151 in the preserved tyrosine kinase region, which contains 50-60% of the phosphate after insulin stimulation (Tornqvist et al., 1988 White et al., 1988b), are crucial for autoactivation (Ellis et al., 1986). 

Like the protein and inositide kinases, the 3D structures of APHs display two distinct domains The N-terminal P-sheet region is responsible for ATP binding, and the a-helical C-terminal provides the aminoglycoside recognition site. The active site, where phosphate transfer occurs, lies at the interface of the two domains. APHs also contain the H-G/N-D-XXXX-N sequence motif, which is common among protein kinases and involved in phosphate transfer catalysis. Structural homology to protein kinases is extended to function, as it has been demonstrated that APHs have weak but measurable protein kinase activity. ... 

Fig. 7.2. Structure and substrate binding sites of Ser/Thr-spedfic protein kinases, a) Peptide binding site structure of the catalytic subunit of the cAMP-dependent protein kinase A from mouse, with bound inhibitor peptide PKI (5-22), shown in dark in the figure. PKI (5-22) is a fragment (amino adds 5-22) of the naturally occurring heat-stable protein kinase inhibitor PKI. The inhibitor peptide binds in the region of the substrate binding site between the two lobes of protein kinase A (Knighton et al., 1991). The P-loop is involved in binding the phosphate residue of ATP. b) ATP binding site structure of casein kinase I with bound Mg-ATP. The Mg is shown as a sphere. MOLSKRIPT representation according to Kraulis, (1991).

FIGURE 2—33. Gene regulation by neurotransmitters, part 3. Once activated, protein kinase phos-phorylates a transcription factor (TF). Attaching phosphate (P04) to this transcription factor activates it so it can bind to the regulatory region of a gene. 

The EGF receptor possesses intrinsic kinase activity and will transfer the terminal phosphate of ATP specifically to the parahydroxyl group of tyrosine residues in certain substrate proteins. The region that encodes this enzyme activity lies between residues 690 and 940 of the cytoplasmic domain (Fig. 1). These sequences are the central, common structural feature shared by the GFRs which possess ligand-stimulated tyrosine kinase activity. Towards the N-terminal end of this region are residues involved in binding the substrate ATP, but it is not known which residues interact with substrate proteins. This is the only known activity of the EGF... 

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