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Reversible pH-controlled DNA-binding peptide nanotweezers: an in-silico study.

Sharma G, Rege K, Budil DE, Yarmush ML, Mavroidis C - Int J Nanomedicine (2008)

Bottom Line: Modulating the solution pH between neutral and acidic values results in the reversible movement of helices toward and away from each other and creates a complete closed-open-closed transition cycle between the helices.The efficacy of the mutant that demonstrated the most significant reversible actuation for environmentally responsive modulation of DNA-binding activity was also demonstrated.Our results have significant implications in bioseparations and in the engineering of novel transcription factors.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical and Industrial Engineering, Northeastern University, Boston, MA 02115, USA.

ABSTRACT
We describe the molecular dynamics (MD)-aided engineering design of mutant peptides based on the alpha-helical coiled-coil GCN4 leucine zipper peptide (GCN4-p1) in order to obtain environmentally-responsive nanotweezers. The actuation mechanism of the nanotweezers depends on the modification of electrostatic charges on the residues along the length of the coiled coil. Modulating the solution pH between neutral and acidic values results in the reversible movement of helices toward and away from each other and creates a complete closed-open-closed transition cycle between the helices. Our results indicate that the mutants show a reversible opening of up to 15 A (1.5 nm; approximately 150% of the initial separation) upon pH actuation. Investigation on the physicochemical phenomena that influence conformational properties, structural stability, and reversibility of the coiled-coil peptide-based nanotweezers revealed that a rationale- and design-based approach is needed to engineer stable peptide or macromolecules into stimuli-responsive devices. The efficacy of the mutant that demonstrated the most significant reversible actuation for environmentally responsive modulation of DNA-binding activity was also demonstrated. Our results have significant implications in bioseparations and in the engineering of novel transcription factors.

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a) Structural details of the M3 mutant. The position of                        histidine mutations are shown as spheres. Hydrophobic interactions are shown                        as gray dotted spheres. Note the strong hydrophobic core in the lower half                        of the protein which serves as a ‘hinge’ for the                        nanotweezer design; b) Snapshots of mutant M3 at various time                        instances during a 4 ns simulation. Location of the His246 residue between                        which the opening is measured is shown as a sphere; c) Opening                        dynamics of M3 at low pH. The initial separation between the two chains was                        11 Å which gradually increased to 27 Å at the 2.5 ns                        stage and then stabilized for the rest of the simulation giving a net                        opening of 16 Å; d) Evolution of the secondary                        structure elements of M3 mutant as a function of time. The structural                        features of the two chains were conserved throughout the simulation.
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f5-ijn-3-505: a) Structural details of the M3 mutant. The position of histidine mutations are shown as spheres. Hydrophobic interactions are shown as gray dotted spheres. Note the strong hydrophobic core in the lower half of the protein which serves as a ‘hinge’ for the nanotweezer design; b) Snapshots of mutant M3 at various time instances during a 4 ns simulation. Location of the His246 residue between which the opening is measured is shown as a sphere; c) Opening dynamics of M3 at low pH. The initial separation between the two chains was 11 Å which gradually increased to 27 Å at the 2.5 ns stage and then stabilized for the rest of the simulation giving a net opening of 16 Å; d) Evolution of the secondary structure elements of M3 mutant as a function of time. The structural features of the two chains were conserved throughout the simulation.

Mentions: Mutant M3 was designed next with the following point mutations in addition to the N-terminal histidine tag: L253H, K256H, E259H, L261H, Y265H. These mutation sites correspond to the d, g, c, e, and b positions respectively on the helical wheel diagram (Figure 1c). Figure 5a shows the starting structure of M3 with the His mutation sites shown as spheres. M3 has a uniform distribution of His residues along the helical chain which results in a spatial distribution of electrostatic charges in addition to the concentrated charges from the distal His-tags. Further, the L253H, L261H, and Y265H mutations replace the hydrophobic leucine and tyrosine residues with polar His residues thereby significantly reducing the strength of the hydrophobic interactions towards the N-terminal and ‘middle’ regions of the coiled-coil core while maintaining the strong hydrophobic core in the C-terminal region. This evolved design was therefore a balance between repulsive forces that can induce the actuation mechanism at low pH and strong hydrophobic interactions that can (i) maintain the coiled-coil structure and (ii) serve as the restoring force for the ‘hinge’ action in order to restore the original conformation of the peptide at neutral pH. We verified, as predicted from its MultiCoil score (Wolf et al 1997), that these mutations do not destabilize the coiled-coil formation propensity of the individual helices. Interestingly, of all the mutants evaluated, M3 demonstrated the highest probability (0.9) of dimer formation in solution (Figure S-1).


Reversible pH-controlled DNA-binding peptide nanotweezers: an in-silico study.

Sharma G, Rege K, Budil DE, Yarmush ML, Mavroidis C - Int J Nanomedicine (2008)

a) Structural details of the M3 mutant. The position of                        histidine mutations are shown as spheres. Hydrophobic interactions are shown                        as gray dotted spheres. Note the strong hydrophobic core in the lower half                        of the protein which serves as a ‘hinge’ for the                        nanotweezer design; b) Snapshots of mutant M3 at various time                        instances during a 4 ns simulation. Location of the His246 residue between                        which the opening is measured is shown as a sphere; c) Opening                        dynamics of M3 at low pH. The initial separation between the two chains was                        11 Å which gradually increased to 27 Å at the 2.5 ns                        stage and then stabilized for the rest of the simulation giving a net                        opening of 16 Å; d) Evolution of the secondary                        structure elements of M3 mutant as a function of time. The structural                        features of the two chains were conserved throughout the simulation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2636583&req=5

f5-ijn-3-505: a) Structural details of the M3 mutant. The position of histidine mutations are shown as spheres. Hydrophobic interactions are shown as gray dotted spheres. Note the strong hydrophobic core in the lower half of the protein which serves as a ‘hinge’ for the nanotweezer design; b) Snapshots of mutant M3 at various time instances during a 4 ns simulation. Location of the His246 residue between which the opening is measured is shown as a sphere; c) Opening dynamics of M3 at low pH. The initial separation between the two chains was 11 Å which gradually increased to 27 Å at the 2.5 ns stage and then stabilized for the rest of the simulation giving a net opening of 16 Å; d) Evolution of the secondary structure elements of M3 mutant as a function of time. The structural features of the two chains were conserved throughout the simulation.
Mentions: Mutant M3 was designed next with the following point mutations in addition to the N-terminal histidine tag: L253H, K256H, E259H, L261H, Y265H. These mutation sites correspond to the d, g, c, e, and b positions respectively on the helical wheel diagram (Figure 1c). Figure 5a shows the starting structure of M3 with the His mutation sites shown as spheres. M3 has a uniform distribution of His residues along the helical chain which results in a spatial distribution of electrostatic charges in addition to the concentrated charges from the distal His-tags. Further, the L253H, L261H, and Y265H mutations replace the hydrophobic leucine and tyrosine residues with polar His residues thereby significantly reducing the strength of the hydrophobic interactions towards the N-terminal and ‘middle’ regions of the coiled-coil core while maintaining the strong hydrophobic core in the C-terminal region. This evolved design was therefore a balance between repulsive forces that can induce the actuation mechanism at low pH and strong hydrophobic interactions that can (i) maintain the coiled-coil structure and (ii) serve as the restoring force for the ‘hinge’ action in order to restore the original conformation of the peptide at neutral pH. We verified, as predicted from its MultiCoil score (Wolf et al 1997), that these mutations do not destabilize the coiled-coil formation propensity of the individual helices. Interestingly, of all the mutants evaluated, M3 demonstrated the highest probability (0.9) of dimer formation in solution (Figure S-1).

Bottom Line: Modulating the solution pH between neutral and acidic values results in the reversible movement of helices toward and away from each other and creates a complete closed-open-closed transition cycle between the helices.The efficacy of the mutant that demonstrated the most significant reversible actuation for environmentally responsive modulation of DNA-binding activity was also demonstrated.Our results have significant implications in bioseparations and in the engineering of novel transcription factors.

View Article: PubMed Central - PubMed

Affiliation: Department of Mechanical and Industrial Engineering, Northeastern University, Boston, MA 02115, USA.

ABSTRACT
We describe the molecular dynamics (MD)-aided engineering design of mutant peptides based on the alpha-helical coiled-coil GCN4 leucine zipper peptide (GCN4-p1) in order to obtain environmentally-responsive nanotweezers. The actuation mechanism of the nanotweezers depends on the modification of electrostatic charges on the residues along the length of the coiled coil. Modulating the solution pH between neutral and acidic values results in the reversible movement of helices toward and away from each other and creates a complete closed-open-closed transition cycle between the helices. Our results indicate that the mutants show a reversible opening of up to 15 A (1.5 nm; approximately 150% of the initial separation) upon pH actuation. Investigation on the physicochemical phenomena that influence conformational properties, structural stability, and reversibility of the coiled-coil peptide-based nanotweezers revealed that a rationale- and design-based approach is needed to engineer stable peptide or macromolecules into stimuli-responsive devices. The efficacy of the mutant that demonstrated the most significant reversible actuation for environmentally responsive modulation of DNA-binding activity was also demonstrated. Our results have significant implications in bioseparations and in the engineering of novel transcription factors.

Show MeSH
Related in: MedlinePlus