Fig 1.
Comparative sequence and structural analysis.
(A) Multiple sequence aligment of group-I. (B) Multiple sequence aligment of group-II. (C) Superposed ribbon diagram PAKs group-I members. (D) Superposed ribbon diagram PAKs group-II members. Secondary structure is delineated at the upper side of sequence as β-strands (arrows) and α-helices (coil). ATP-binding residue (β3) Lys, (αC) Glu, (β5) Glu and (β8) Lys are delineated in navy blue, brown, khaki, magenta and aqumarine colors, respectively. Activation loop (T-loop) and phosphorylated residues (T/S) are delineated in lime green and sandy brown colors. N- and C-lobes are indicated in light blue and white colors, respectively.
Fig 2.
MD simulation analysis of group-I PAKs.
(A and B) RMSD and RMSF plots for PAK1 (golden) and PAK1Tpo423 (red), (C and D) RMSD and RMSF plots for PAK2 (dark green) and PAK2Tpo402 (purple) and (E and F) RMSD and RMSF plots for PAK3 (sky blue) and PAK3Tpo436 (dark blue), respectively. (G) RMSF values of highly fluctuating residues are plotted and indicated by the corresponding colors.
Fig 3.
Comparative ribbon analysis of group-I PAKs on the basis of MD simulation analysis.
Ribbon representation of (A and B) PAK1 and PAK1Tpo423, (C and D) PAK2 and PAK2Tpo402 and (E and F) PAK3 and PAK3Tpo436 structures. Ribbon view color code: N-lobe, C-lobe and activation loop are shown in light blue, white and lime green colors, respectively. Gly-loop is shown in orange red color, while phosphorylated (Tpo423,Tpo402 and Tpo436) and unphosporylated (Thr423, Thr402 and Thr436) residues are shown in sandy brown sphere. β5-specific Glu and β8-specific Lys residues are shown in magenta and aqumarine colors, respectively. The residual distances are highlighted by black dotted lines. These color codes are used for all the ribbon view representation for group-I and group-II PAKs structures.
Fig 4.
Comparative surface view analysis of group-I PAKs on the basis of MD simulation.
Surface view of (A and B) PAK1 and PAK1Tpo423, (C and D) inactive and active PAK1, rotated at 180°, (E and F) PAK2 and PAK2Tpo402, (G and H) inactive and active PAK2, rotated at 180°, (I and J) PAK3 and PAK3Tpo436, (K and L) inactive and active PAK3, rotated at 180°. Surface view color code: C-lobe and activation loop are shown in light blue, white and lime green colors, respectively. Gly-loop is shown in orange red color. β6, β7 and β8-strands are shown in deep pink colors, respectively. These color codes are used for all the Surface view representation for group-I and group-II PAKs structures.
Fig 5.
RMSD, RMSF and fluctuated residue plots for to explore the fluctuation and stability of PAKs group-II.
(A and B) RMSD and RMSF plots for PAK4 (light green) and PAK4sep474 (maroon), (C and D) RMSD and RMSF plots for PAK5 (purple) and PAK5Sep602 (deep pink), and (E and F) RMSD and RMSF plots for PAK6 (golden rod) and PAK3Sep560 (dark green), respectively. (G) RMSF values of highly Fluctuating residues are plotted and indicated by the corresponding colors.
Fig 6.
Comparative ribbon analysis of group-II PAKs on the basis of MD simulation analysis.
Ribbon representation of (A and B) PAK4 and PAK4Sep474, (C and D) PAK5 and PAK5Sep602 and (E and F) PAK6 and PAK6Sep560 structures. Color codes are given in the Fig 3 legend.
Fig 7.
Comparative surface view analysis of group-II PAKs on the basis of MD simulation.
Surface view of (A and B) PAK4 and PAK4Sep474, (C and D) inactive and active PAK4, rotated at 180°, (E and F) PAK5 and PAK5Sep602, (G and H) inactive and active PAK5, rotated at 180°, (I and J) PAK6 and PAK6Sep560, (K and L) inactive and active PAK6, rotated at 180°. Color codes are given in the Fig 4 legend.
Fig 8.
(a) (5-Phenyl-1,2-oxazol-3-yl)[7-{[4-(2-pyridinyl)-1-piperazinyl]methyl}-2,3-dihydro-1,4-benzoxazepin-4(5H)-yl]methanone, (b) 2-{2-[7-(5,8-Dimethoxy-4-methyl-2-quinolinyl)-9-methoxy-2,3-dihydro-1,4-benzoxazepin-4(5H)-yl]-2-oxoethyl}-4-methyl-1(2H)-phthalazinone, (c) N-({2-[3-(Benzoylamino)phenyl]-5-methyl-1,3-oxazol-4-yl}methyl)-3-(2-oxo-1-imidazolidinyl)benzamide, (d) 2-(1H-Indol-3-yl)-1-{4-[2-(tetrahydro-2-furanyl)-5-(trifluoromethyl)-1H-benzimidazol-1-yl]-1-piperidinyl}ethanone, (e) Methyl 2-[(4-fluoro-2-methoxybenzyl)amino]isonicotinate, (f) N-{[7-(3-Cyclohexen-1-ylcarbonyl)-3-methyl-5,6,7,8-tetrahydro-2,7-naphthyridin-4-yl]methyl}-2-methyl-3-furamide, (g) 1-({2-[2-(Ethylamino)-5-pyrimidinyl]-6-fluoro-4-quinolinyl}carbonyl)-3-piperidinecarbonitrile, (h) 2-[4-(8-Methoxy-2-methyl-5-quinolinyl)-1H-pyrazol-1-yl]-N-(2-methyl-2H-1,2,3-triazol-4-yl)acetamide, (i) 1-(1,5-Dimethyl-1H-1,2,4-triazol-3-yl)-3-[2-(4-methoxyphenyl)ethyl]urea, (j) 2-{4-[(1S,4S)-2-Azabicyclo[2.2.1]hept-2-ylmethyl]phenyl}-6-(3-pyridinyl)-4(1H)-pyrimidinone, (k) N-[2-(5-Fluoro-1H-benzimidazol-2-yl)ethyl]-5-methyl-2-pyrazinecarboxamide and (l) (2E)-N-{[7-Chloro-5-(2,5-dimethoxyphenyl)-2,3-dihydro-1-benzofuran-2-yl]methyl}-3-(2-pyridinyl)acrylamide.
Fig 9.
Comparative binding mode analysis of proposed inhibitors.
(A) PAK1Tpo423, (B) PAK1, (C) PAK4Sep474 and (D) PAK4. The inhibitors (a-l) are shown in orange, nave blue, chartreuse, orchid, dark red, yellow, dark state gray, aquamarine, coral, golden rod, rosy brown and dark magenta colors, respectively.
Table 1.
Comparative binding energy values of inhibitor-bound PAK1 and PAK4 (active and inactive states).
Fig 10.
Moleculer mechanistic insights into functional characteristics of PAK family.
PAKs are activated in response to extracellular stimuli and recruit diverse signaling pathways. PAKs are now known to be potential regulators of intracellular activity, cytoskeleton remodeling, cell survival, transformation, cell cycle and gene transcription pathways [57]. PAK2 activation by Rac, cdc42 cleavage, caspases or caspase-like proteases correlates with the programmed cell death. Thus, PAK2 is unique among the PAK isoforms due to its involvement in the stimulation of cell survival and cell death events depending on the activation mechanism [58]. Group II PAK signaling pathways have been observed downstream to membrane receptors and multiple potential regulators of intracellular activity. The three family members appear to have distinct and overlapping cellular functions and interact with an array of downstream effectors to elicit their cellular responses. EGF, epidermal growth factor; HGF, hepatocyte growth factor; PP1B, protein phosphatase 1B [21].