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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) Schematic of the operating principle of nanotweezer. Initial                        ‘closed’ state at neutral pH                        (Left). Final ‘open’ state                        generated at low pH (Right). The plus                        signs in the ‘open’ state represent the location of                        engineered histidine residues which becomes positively charged at low pH                        thereby creating electrostatic repulsive forces; b) Nanotweezer                        mutants; Wild-Type (WT), Mutants M1, M2, M3, and the M3 control (M3CT). The                        position of glycine tag in WT and M3CT is shown in                        ‘bond’ representation. Position of His-tags and                        histidine mutations in other mutants is shown in dark shades.
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f2-ijn-3-505: a) Schematic of the operating principle of nanotweezer. Initial ‘closed’ state at neutral pH (Left). Final ‘open’ state generated at low pH (Right). The plus signs in the ‘open’ state represent the location of engineered histidine residues which becomes positively charged at low pH thereby creating electrostatic repulsive forces; b) Nanotweezer mutants; Wild-Type (WT), Mutants M1, M2, M3, and the M3 control (M3CT). The position of glycine tag in WT and M3CT is shown in ‘bond’ representation. Position of His-tags and histidine mutations in other mutants is shown in dark shades.

Mentions: In the present work, we describe the molecular dynamics-aided design, concept evolution and biophysical characterization of an engineered peptide nanotweezer based on the coiled-coil GCN4-LZ. The simplicity, regularity in structural organization, and availability of the peptide crystal structure allows the engineering of GCN4-LZ to develop functional nanoscale elements. The GCN4-LZ was engineered to obtain pH-dependent nanotweezers involving the lateral displacement of the two helices relative to each other. The reversible actuation mechanism depends on the generation of similar electrostatic charges along the peptide chain which forces the two coils to repel each other, creating a closed-to-open transition. Neutralizing these charges leads to an open-to-closed transition and restitution of the original structure stabilized primarily by hydrophobic interactions. Figure 2a shows a schematic of the nanotweezer operating principle. A broader impact of this study was the analysis of coiled-coil stability under different pH conditions in addition to an in-depth investigation into the effect of point mutations and electrostatic forces on coiled-coil secondary structure. Based on these studies, we propose the design of a DNA-binding modulator element based on the pH-driven nanotweezer architecture and show preliminary simulation results to support our hypothesis. The development of such a DNA-binding modulator has implications in transcription factor engineering wherein one of the focuses is the construction of designer transcription factors for various therapeutic and research applications (Beerli and Barbas 2002). We also describe the development of key design principles required for incorporating flexibility in rigid peptide motifs which can have implications in computational drug design (Carlson and McCammon 2000), design of protein-based biosensors (Gooding et al 2003) and molecular motors (Sun et al 2003).


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) Schematic of the operating principle of nanotweezer. Initial                        ‘closed’ state at neutral pH                        (Left). Final ‘open’ state                        generated at low pH (Right). The plus                        signs in the ‘open’ state represent the location of                        engineered histidine residues which becomes positively charged at low pH                        thereby creating electrostatic repulsive forces; b) Nanotweezer                        mutants; Wild-Type (WT), Mutants M1, M2, M3, and the M3 control (M3CT). The                        position of glycine tag in WT and M3CT is shown in                        ‘bond’ representation. Position of His-tags and                        histidine mutations in other mutants is shown in dark shades.
© Copyright Policy
Related In: Results  -  Collection

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

f2-ijn-3-505: a) Schematic of the operating principle of nanotweezer. Initial ‘closed’ state at neutral pH (Left). Final ‘open’ state generated at low pH (Right). The plus signs in the ‘open’ state represent the location of engineered histidine residues which becomes positively charged at low pH thereby creating electrostatic repulsive forces; b) Nanotweezer mutants; Wild-Type (WT), Mutants M1, M2, M3, and the M3 control (M3CT). The position of glycine tag in WT and M3CT is shown in ‘bond’ representation. Position of His-tags and histidine mutations in other mutants is shown in dark shades.
Mentions: In the present work, we describe the molecular dynamics-aided design, concept evolution and biophysical characterization of an engineered peptide nanotweezer based on the coiled-coil GCN4-LZ. The simplicity, regularity in structural organization, and availability of the peptide crystal structure allows the engineering of GCN4-LZ to develop functional nanoscale elements. The GCN4-LZ was engineered to obtain pH-dependent nanotweezers involving the lateral displacement of the two helices relative to each other. The reversible actuation mechanism depends on the generation of similar electrostatic charges along the peptide chain which forces the two coils to repel each other, creating a closed-to-open transition. Neutralizing these charges leads to an open-to-closed transition and restitution of the original structure stabilized primarily by hydrophobic interactions. Figure 2a shows a schematic of the nanotweezer operating principle. A broader impact of this study was the analysis of coiled-coil stability under different pH conditions in addition to an in-depth investigation into the effect of point mutations and electrostatic forces on coiled-coil secondary structure. Based on these studies, we propose the design of a DNA-binding modulator element based on the pH-driven nanotweezer architecture and show preliminary simulation results to support our hypothesis. The development of such a DNA-binding modulator has implications in transcription factor engineering wherein one of the focuses is the construction of designer transcription factors for various therapeutic and research applications (Beerli and Barbas 2002). We also describe the development of key design principles required for incorporating flexibility in rigid peptide motifs which can have implications in computational drug design (Carlson and McCammon 2000), design of protein-based biosensors (Gooding et al 2003) and molecular motors (Sun et al 2003).

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