KinCoRe
Kinase Conformation Resource
A web resource for protein kinase sequence, structure and phylogeny.
Kinase Conformation Resource
A web resource for protein kinase sequence, structure and phylogeny.
Protein kinases (PKs) are enzymes that transfer phosphoryl group from an ATP molecule to Ser, Thr or Tyr residue of the substrate protein. The human genome consists 484 PK genes (497 domains) that are divided broadly into nine families based on their sequences namely, AGC, CAMK, CK1, CMGC, NEK, RGC, STE, TKL, TYR and OTHER (unclassified). They share a conserved structural fold consisting of two lobes: an N-terminal lobe, formed by five stranded β-sheet with an α-helix called the C-helix, and a C-terminal lobe comprising six α-helices. The two lobes are connected by a flexible region in the middle which forms the ATP binding active site of the protein.
The activation loop is typically 20-30 residues in length and is the most critical secondary structural element of the active site of PKs. It is in completely extended conformation in the catalytically active state of the enzyme facilitating the binding of ATP molecule and the substrate. However, it folds on the surface of the protein in different kinds of inactive states. The activation loop begins with a conserved sequence motif called DFGmotif (Asp, Phe, and Gly residues). These residues are observed to be in a unique orientation when the loop is extended (active state) but display remarkable flexibility in folded (inactive) loop conformations.
Typical structure of protein kinase
Multiple orientations of DFG-Phe in EGFR
We have determined the location of DFG-Phe ring in the binding pocket based on its distance from two
conserved residues,
Based on the spatial location of DFG-Phe ring in the binding pocket we have classified kinase structures into three broad groups:
Each spatial group consists of multiple closely related conformations. To cluster these conformations we used the backbone dihedrals (φ,ψ) of X-DFG (residue before conserved Asp), DFG-Asp, DFG-Phe and side chain dihedral (χ1) of DFG-Phe. These dihedrals we used to compute a distance matrix which is then provided as an input to DBSCAN (Density-based spatial clustering of applications with noise), a density-based clustering algorithm. The different clusters observed are labeled on the basis of Ramachandran region (A, B, L, and E) occupied by XDF residue backbone and the DFG-Phe χ1 rotamer (minus = -60°; plus = +60°; trans = 180°).
For the DFGin group we obtained six clusters labeled as BLAminus, BLAplus, ABAminus, BLBminus, BLBplus, and BLBtrans. All the catalytically primed structures (ATP+Mg bound and activation loop phosphorylated) are observed in the BLAminus cluster.
For the DFGout group we obtained just one cluster. In this cluster, the X-D-F residues occupy the B-B-A regions of the Ramachandran map and DFG-Phe is in a -60° rotamer. More than 82% of Type 2 inhibitor bound structures are BBAminus; the remainder are in the DFGout noise group.
The structures in the DFGinter conformation display more variability than the other states. For the DFGinter group we obtained only one small cluster. The X-D-F residues are in a B-A-B conformation and the DFG-Phe residue is observed in a trans rotamer with a few chains displaying a rotamer orientation between g-minus and trans.
A. Pairwise comparisons of 5 different states of human BRAF kinase
DFGin-BLBplus structures (“SRC-inactive” conformation in blue with grayish-blue activation loop)
compared with DFGin-BLAminus (orange, active conformations), BLBminus (magenta-pink), DFGinBLAplus (“FGFR-inactive”, cyan-lightcyan), and DFGout-BBAminus (Type-2 ligand binding, lightbrownyellow). A small number of outlier structures in some classes are not shown. In each case, the position of the activation loop and the C-helix is correlated with
the state of the kinase, and differs among the 5 states.
B. BLAminus and BLBplus states of EGFR
EGFR BLAminus (107
chains, left, orange) and BLBplus (79 chains, right, blue) structures with both the activation loop
(right side of each image) and C-terminal tail (left side of each image) shown in lighter colors. The C-terminus of each group is shown in magenta spheres. The state of the activation loop is highly correlated with the
position of the C-terminal tail. In the active BLAminus structures, the tail is mostly coil and reaches up
to strands B2 and B3 of the N-terminal domain. In the “SRC-inactive” BLBplus state, the tail contains a
helix (residues 993-1002) in contact with the C-terminal domain, and then turns around with the C-terminus in contact with the I-helix of the C-terminal domain.
C. Results of searches with the Kincore website
ATP-bound BLAminus structures from 14
different kinases in 6 kinase families (left) and BBAminus structures from 29 kinases with bound
Type 2 inhibitors (right).In the
BLAminus structures, the position of the DFG Phe (in yellow) and the conformation of the DFGmotif at
the beginning of the activation loop (shown in magenta) and the overall position of the activation loop
are consistent across the structures.
We have classified PK inhibitors into five groups based on the region of protein they bind to.
A list of FDA approved PK inhibitors with known structures can be accessed here.