Abstract
BackgroundProtein kinases (PKs) have emerged as the largest family of signaling proteins in eukaryotic cells and are involved in every aspect of cellular regulation. Great progresses have been made in understanding the mechanisms of PKs phosphorylating their substrates, but the detailed mechanisms, by which PKs ensure their substrate specificity with their structurally conserved catalytic domains, still have not been adequately understood. Correlated mutation analysis based on large sets of diverse sequence data may provide new insights into this question.Methodology/Principal FindingsStatistical coupling, residue correlation and mutual information analyses along with clustering were applied to analyze the structure-based multiple sequence alignment of the catalytic domains of the Ser/Thr PK family. Two clusters of highly coupled sites were identified. Mapping these positions onto the 3D structure of PK catalytic domain showed that these two groups of positions form two physically close networks. We named these two networks as θ-shaped and γ-shaped networks, respectively.Conclusions/SignificanceThe θ-shaped network links the active site cleft and the substrate binding regions, and might participate in PKs recognizing and interacting with their substrates. The γ-shaped network is mainly situated in one side of substrate binding regions, linking the activation loop and the substrate binding regions. It might play a role in supporting the activation loop and substrate binding regions before catalysis, and participate in product releasing after phosphoryl transfer. Our results exhibit significant correlations with experimental observations, and can be used as a guide to further experimental and theoretical studies on the mechanisms of PKs interacting with their substrates.
Highlights
Phosphorylation of protein substrates by Protein kinases (PKs) is the most abundant and important type of cellular regulation [1]
Sequence Collection and Pretreatment In order to ensure that sequences of alignment are representative and diverse, we collected the homologue sequences by using seventeen different sequences as initial query sequences. These initial query sequences come from nine eukaryotic organisms including vertebrate, invertebrate, plant and fungus, and they are distributed over all subfamilies of Ser/Thr PKs (See Supporting Information Figure S1) [8,28]
These collected homologue sequences are from 206 eukaryotic organisms, and can adequately represent the properties of the Ser/Thr PK catalytic domain family and eliminate the phylogenetic bias in the collection of sequences
Summary
Phosphorylation of protein substrates by PKs is the most abundant and important type of cellular regulation [1]. The vast majority of PKs are Ser/Thr PKs. Previous studies [2,3] on PK structures have shown that the basic fold of the catalytic domains of PKs is structurally well conserved (i.e., two-lobe structure), and the peptide substrates are always held in the groove between the two lobes in many PK-substrate complex structures. Great progresses have been made in understanding the mechanisms of PKs phosphorylating their proper substrates [6,7] These mechanisms include the structure of catalytic cleft, consensus sequences, local and distal interactions between kinase and substrate. Great progresses have been made in understanding the mechanisms of PKs phosphorylating their substrates, but the detailed mechanisms, by which PKs ensure their substrate specificity with their structurally conserved catalytic domains, still have not been adequately understood. Correlated mutation analysis based on large sets of diverse sequence data may provide new insights into this question
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