Investigation into Early Steps of Actin Recognition by the Intrinsically Disordered N-WASP Domain V

Figures

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  Figure 1. Probability of the N-WASP domain V residues to be distant by less than 4 Å from actin. Orange and magenta dashed lines indicate the protein regions in α-helix (as revealed by the X-ray structure 2VCP [27]) and the consensus sequences “LKKV” [14,31], respectively.

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  Figure 2. (A) Probability of actin residues to be distant by less than 4 Å from domain V regions 9–18 or 37–46. Red dashed lines indicate actin residues in contact with N-WASP helical segment in structure 2VCP [27]. (B) Views of actin’s surface colored proportionally to previous probabilities. Blue, white, and red colors indicate actin residues with low, intermediate, and high probabilities to be contacted by domain V, respectively. As a reference, yellow and green ribbons represent the second WH2 motif helical region and conserved sequence LKSV as observed in 2VCP [27].

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  Figure 3. Side view of the two best 1:1 actin–domain V encounter complexes with N-WASP segment 9–18 (left) or 37–46 (right) located and oriented as in structure 2VCP. Black balls are N-terminal Cα-atoms of domain V. Red and magenta ribbons represent its regions 9–18 or 37–46 and consensus sequences “LKKV”, respectively. As a reference, yellow and green ribbons indicate the helical and 50LKSV53 regions of domain VC in 2VCP.

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  Figure 4. Time evolutions of RMSD relative to structure 2VCP, after fitting MD trajectories on crystallographic actin, for actin (black) and segments 9–18 (orange) and 37–46 (cyan) of N-WASP domain V.

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  Figure 5. Time evolutions of minimal distance to actin of segments 22LKKV25 (black) and 50LKSV53 (brown) of N-WASP domain V. RMSD relative to structure 2VCP, after fitting trajectories on actin, are also displayed as a function of time for segments 22LKKV25 (orange) and 50LKSV53 (cyan).

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  Figure 6. (A) Probability of actin residues to be distant by less than 4 Å from N-WASP segments 22LKKV25 or 50LKSV53 in CplxA_MD1 (red), CplxA_MD2 (orange), CplxB_MD1 (cyan), and CplxB_MD2 (blue). Brown dashed lines indicate the actin residues (G23, D24, D25, R28, and S344) in contact with N-WASP segment 50LKSV53 in structure 2VCP [27]). (B–D) Front views of the actin’s surface colored proportionally to the previous probabilities. Red, orange, and blue colors indicate actin residues with high probabilities to be contacted by N-WASP segments 22LKKV25 or 50LKSV53 in simulations CplxA_MD1 (B), CplxA_MD2 (C), and CplxB_MD2 (D), respectively. As a reference, yellow and green ribbons represent the helical region and the conserved sequence LKSV of the second WH2 motif observed in structure 2VCP [27].

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  Figure 7. Time evolutions of minimal distances between actin and segment 28NSRPVS33 in simulations of CplxA (red and orange lines) and segment 56GQESTP61 in simulations of CplxB (cyan and blue lines). For comparison, time evolutions of minimal distances between actin and segments 22LKKV25 and 50LKSV53 are displayed with black and brown lines, respectively.

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  Figure 8. Actin residues distant by less than 4 Å from N-WASP segments 28NSRPVS33 or 56GQESTP61 in CplxA_MD1 (red), CplxA_MD2 (orange), CplxB_MD1 (cyan), and CplxB_MD2 (blue). As a reference, yellow and green ribbons represent the helical region and the conserved sequence LKSV of the second WH2 motif observed in structure 2VCP [27].

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  Figure 9. Probability of the N-WASP domain V residues to be distant by less than 4 Å from actin dimer. Orange and magenta dashed lines indicate the N-WASP regions in α-helix (as revealed by the X-ray structure 2VCP [27]) and the consensus sequences “LKKV” [14,31], respectively.

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  Figure 10. (A) Residue-specific probability of actin dimer chain A (bottom) and chain B (top) to be distant by less than 4 Å from N-WASP domain V regions 9–18 or 37–46 in the ensemble of 7540 ternary complexes generated by docking. Red dashed lines indicate the actin residues in contact with the N-WASP helical segment in the X-ray structure 2VCP [27]. (B,C) Back and front views of the actin dimer surface colored proportionally to the previous probabilities. Blue, white, and red colors indicate actin residues with low, intermediate, and high probabilities to be contacted by N-WASP domain V regions 9–18 or 37–46, respectively. As a reference, yellow and green ribbons represent helical regions and consensus sequences “LKKV” of the two WH2 motifs observed in the X-ray structure 3M3N [26].

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  Figure 11. Side view of the two best 2:1 actin–domain V encounter complexes with N-WASP segment 9–18 (left) or 37–46 (right) located and oriented as in structure 3M3N. Black balls are N-terminal Cα-atoms of domain V. Red and magenta ribbons represent its regions 9–18 or 37–46 and consensus sequences “LKKV”, respectively. As a reference, yellow and green ribbons indicate the helical and LKKV regions of domain V in 3M3N.

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  Figure 12. Possible pathways toward the formation of a 2:1 actin–domain V complex with a longitudinal actin dimer as observed in 3M3N. Starting from two actin chains and one N-WASP domain V (State I), three possible binary encounter complexes can be formed (States II-a, II-b, and II-c), leading to three possible intermediate ternary complexes (States III-a, III-b, and III-c) just before the final structure (State IV). Cyan and red arrows indicate the binding of the N-WASP first and second WH2 motif to actin chain A and B, respectively. Dark grey arrows represent the binding of two actins into a longitudinal dimer.