![]() ![]() Thr 402 and Ser 141 are critical for Pak2 regulation and activation. Cleavage by caspase 3 or binding of Cdc42(GTP) facilitates autophosphorylation at Ser 141, Ser 165 and Thr 402. Activation of the protein kinase requires autophosphorylation at seven serine sites in the regulatory domain and one conserved threonine in the catalytic domain. Pak2 has a basal autophosphorylation activity wherein five serines in the regulatory domain of Pak2 can be autophosphorylated. Pak2 is primarily inactive during growth, and is transiently activated by Cdc42 in response to moderate stress, and constitutively activated during apoptosis by caspase cleavage. Pak2 belongs to the serine/threonine protein kinase family containing the conventional Pak 1–3 and Pak 4–6. These reciprocally coupled pairs of residues are the structurally or functionally coupled residues occurring through evolution. The distilled results of the statistical coupling analysis eliminated all perturbations from a single direction. We incorporated this concept and retrieved all reciprocal coupling residues of Pak2. The correlation pattern showed the residues coupled reciprocally to structural determinants of the G protein-coupled receptors. used matrix clustering analysis to find the correlation patterns from the two-dimensional SCA array. The SCA results are presented in a two-dimensional array (matrix), representing the correlation values between any of the two residues. Cross-correlation analysis of sequence-based SCA and structure-based molecular dynamics predicted energetic coupling residues essential for HhaI ( Haemophilus haemolyticus I) methyltransferase catalysis. Other research applied the matrix clustering analysis to systematically analyze large amounts of SCA data to identify the key residues for the structures of the G protein-coupled receptors, G proteins, and RXR heterodimers. The statistical coupling through sequence-based analysis was introduced as a reporter of thermodynamic coupling in proteins with PDZ domains. This analysis retrieves two co-evolved residues in a protein for either structural or functional reasons. Statistical coupling analysis (SCA) regards evolution as a natural mutagenesis process and utilizes the known protein sequences to economically examine comprehensive correlations between amino acid residues. This indicates these evolutionary conserved residues are crucial for the catalytic activity of PKA and Pak2. This is the inhibitor (PKI) binding region in PKA and the activation groove for Pak2. Reciprocal coupling residues exposed to the solvent line up as an activation groove. Nine pairs of reciprocal coupling residues crucial for enzymatic activity and structural stabilization were identified. Reciprocal coupling analysis by SCA showed the residues perturbed by Thr 402 and the reciprocal coupling pairs formed a network centered at Trp 427 in the F-helix. When compared to the cAMP-dependent protein kinase (PKA) and Src, the perturbation pattern of threonine phosphorylation in the activation loop of Pak2 is similar to that of PKA, and different from the tyrosine phosphorylation pattern of Src. We collected 482 human protein kinase sequences from the kinome database and globally mapped the evolutionary interactions of the residues in the catalytic domain with Thr 402 by sequence-based statistical coupling analysis (SCA). ![]() Regulation of Pak2 activity involves at least two mechanisms: (i) phosphorylation of the conserved Thr 402 in the activation loop and (ii) interaction of the autoinhibitory domain (AID) with the catalytic domain. ![]()
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