BlockMaster: Partitioning Protein Kinase Structures Using Normal-Mode Analysis

Abstract

Protein kinases are key signaling enzymes which are dysregulated in many health disorders and therefore represent major targets of extensive drug discovery efforts. Their regulation in the cell is exerted via various mechanisms, including control of the 3D conformation of their catalytic domains. We developed a procedure, BlockMaster, for partitioning protein structures into semirigid blocks and flexible regions based on residue-residue correlations calculated from normal modes. BlockMaster provided correct partitioning into domains and subdomains of several test set proteins for which documented expert annotation of subdomains exists. When applied to representative structures of protein kinases, BlockMaster identified semirigid blocks within the traditional N-terminal and C-terminal lobes of the kinase domain. In general, the block regions had elevated helical content and reduced, but significant, coil content compared to the nonblock (flexible) regions. The specificity-determining regions, previously used to derive inhibitory peptides, were found to be more flexible in the tyrosine kinases than in serine/threonine kinases. Two blocks were identified which spanned both lobes. The first, which we termed the "pivot" block, included the alphaC-beta4 loop in the N-terminal lobe and part of the activation loop in the C-terminal lobe and appeared in both the active and inactive conformations of the kinases. The second, which we termed the "loop" block, differed between the active and inactive conformations. In the structures of active kinases, this block included part of the activation loop in the C-terminal lobe and the alphaC helix in the N-terminal lobe, representing a known interaction that stabilizes the active conformation. In the inactive structures, this block included G loop residues instead of the alphaC residues. This novel inactive "loop" block may stabilize the inactive conformation and thus downregulate kinase activity.

Dynamics-Driven Allostery in Protein Kinases

“…This community forms strong communication bonds to the five largest communities and represent a central regulatory “hub” that is in a good agreement with an earlier assessment of the αC-helix as a “Signal integration motif” [25]. Both the “pivot” and the “loop” blocks detected by NMA [87] were included in Community C. Although most of the main-chains of the conserved and regulatory residues in the αC-β4 Loop were included in Community C, its side-chains were associated with the neighboring communities. The side-chain of Leu106 that is a part of the R-spine (RS4) was a member of Community A, the side-chain of Val104 that is a part of the “Shell” (Sh1) was associated with the catalytic Community D, and for Phe102 at the turn in the αC-α4 loop both the main-chain and the side-chain belonged to Community E. This unusual property of the αC-β4 Loop reinforces its role as an important “communication hub” that mediates allosteric signals between the R-spine, the ATP-binding pocket and the C-lobe.…”

“…They performed community analysis of these data based on the Clique Percolation Method [93] and detected a varying number (from 8 to 30) of semi-rigid bodies that consisted of four to eight residues. The number and the structure of these communities differed significantly between kinases and their functional states; however, generally larger and more stable communities are observed in the kinase core surrounding the R-spine and the αC-helix in active kinases, supporting observations made by Shudler and Niv [87]. …”

“…Despite a significant simplification of the system, this model, in combination with Normal Mode Analysis (NMA), provides us with a glimpse into very important features of protein long scale dynamic behavior [86]. Shudler and Niv used this approach to analyze the dynamic properties of four protein kinases in their active and inactive forms: two serine/threonine kinases (RAC-alpha serine/threonine-protein kinase (PKB) and Cyclin-dependent kinase 2 (CDK2)) and two tyrosine kinase (Tyrosine-protein kinase ABL1 (ABL1) and Insulin receptor tyrosine kinase (IRK)) [87]. They found that the kinase core can be partitioned into semi-rigid bodies, termed “blocks”, of about five to eight residues each.…”

Deciphering the Arginine-Binding Preferences at the Substrate-Binding Groove of Ser/Thr Kinases by Computational Surface Mapping

“…Actually, it is only in PKC iota that a unique arrangement of the αC helix results in direct involvement of positions 83 and 84 in Arg binding (Figure 4C). Notably, the interplay between all three acidic residues is probably enabled by the flexibility of the acidic cluster region [65].…”

Protein kinases: evolution of dynamic regulatory proteins

“…The smaller N-lobe is dominated by a five stranded β-sheet, which is coupled to a helical subdomain that typically consists of the C-helix (Figure 1). The αC-β4 Loop is the only part of the N-lobe that is functionally and constitutively anchored to the C-lobe [13, 14]. …”