Distinctive H-(RLDL)4-OH peptide complexes potentiate nanostructure self-assembling in water

ISSN 16076729, Doklady Biochemistry and Biophysics, 2012, Vol. 443, pp. 96–99. © Pleiades Publishing, Ltd., 2012. Published in Russian in Doklady Akademii Nauk, 2012, Vol. 443, No. 4, pp. 507–510.

BIOCHEMISTRY, BIOPHYSICS AND MOLECULAR BIOLOGY

Distinctive H–(RLDL)4–OH Peptide Complexes Potentiate Nanostructure SelfAssembling in Water1 A. V. Danilkovicha,b, E. V. Sobolevc, D. A. Tikhonovc, I. P. Udovichenkoa,b, and Corresponding Member of the RAS V. M. Lipkinb,d Received November 30, 2011

DOI: 10.1134/S160767291202010X 1

It is well known that certain ionic and amyloidlike peptides [1] are capable of selfassembling in water to form the amyloidlike fibrils with nanoscaled width; however, these structures might have several hundred nanometers in length. Earlier, it was shown in vitro that spatial network of filaments is formed during the ionic peptide selfassembling. The fibril structure was shown to consist of the peptide molecules in the βsheet conformation oriented perpendicularly to the longitudinal axis of the filament body [2]. Identifica tion of the factors which may influence peptide mole cule selforganization is a highly demanded task in terms of search for new approaches to develop both preventative and medical treatments for amyloidosis and the diseases accompanied by accumulating the amylome and amyloid deposits in tissues and cells [1] and for designing new biomimetics for medicine, since they have advantages for tissue engineering [2]. It should be noted the crucial details of the peptide mol ecule selfassembling, especially the initial stages of protofilament formation, are not clarified yet. The goal of this work was to examine the conformational characteristics of H⎯(RLDL)4–OH peptide com plexes, which are related to type 2.2.1 structures [3], and to establish the spatial organization of the protofilament. To solve these issues, we employed the method of molecular dynamics (MD) [4] to calculate and study the thermodynamic and conformational features of the βsheet structures of H⎯(RLDL)4–OH 1

peptide in dimer, tetramer, and octamer complexes. Certain conclusions were made regarding the spatial organization of these structures, which can support the peptide molecule selfassembling to filamentous nanostructures in water. METHODS The model H⎯(RLDL)4–OH peptide structure for the monomer in βsheet conformation was created with HyperChem8, and then the spatial structures were docked with HEX6. Molecular dynamics (MD) of the peptide molecules and complexes and data analysis were performed with AMBER v.11 [5]. MD experiments with H⎯(RLDL)4–OH peptide struc tures were conducted in explicit water in octahedral PGUcells (table) with model space of AMBER ff03, at 300 K and 1 bar [6]. Water molecules were modeled with TIP3P parameters [7]. Electrostatics were cut over the 10 Å, while the closer interactions in PGU cell images were accounted by the Ewalds summation [8]. Gibbs free energy (ΔG) was calculated with the method of Poisson–Boltzman (MM/PBSA), which is implemented in AMBER v.11. The secondary struc tures of polypeptides were defined according to Kabsch and Sander recommendations [9], which means that the conformation of aminoacid residue were deter mined by the torsions ϕ (C'–N–Cα–C') and ψ (N– Cα–C'–N) for the peptide backbone atoms. The com parison of aminoacids spatial motility was performed by obtaining the values of relative mean square fluctu ations (RMSF, Å) for the peptide backbone atoms during MD experiment. The procedure of calculating RMSF was realized in PTRAJ of AMBER v.11 [5]. RMSF values were calculated for each amino acid res idue, the mean values were found for every amino acid position in the peptide backbone.

The article was translated by the authors.

a Shemyakin–Ovchinnikov

Institute of Bioorganic Chemistry, Russian Academy of Sciences, Pushchino, Moscow oblast, 142290 Russia b Pushchino State University, Pushchino, 142290 Russia c Institute of Mathematical Problems of Biology, Russian Academy of Sciences, Pushchino, 142290 Russia d Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, GSP V437, Moscow, 117997 Russia

RESULTS AND DISCUSSION Molecular docking of H⎯(RLDL)4–OH peptide complex structures in the βsheet conformation has been obtained, and their atomic 3D coordinates were 96

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Main characteristics of the systems been analyzed at PGU H–(RLDL)4–OH peptide structures

Number of water Peptide concentra molecules tion, mol

Monomer Dimer Tetramer“the leaf” Stable tetramer Octamer

6936 7588 8437 8476 9169

0.016 0.029 0.053 0.053 0.097

Gibbs free energy (MM/PBSA), kcal/mol

ΔGasseml *, kcal/mol

ΔGcomplex **, kcal/mol

–782.00 ∓ 12.95 –1640.04 ∓ 19.08 –3334.97 ∓ 29.05 –3309.46 ∓ 28.13 –6767.02 ∓ 51.71

0 –76.04 –54.88 –29.37 –148.10

0 –76.04 –206.97 –181.46 –511.03

* Gibbs free energy of the complex and the sum of the forming parts dimer > tetramer > octamer.

** Gcomplex shows how the Gibbs free energy of the complex differs from the sum of energies for equivalent number of peptide monomers.

stored in corresponding PDB files. Conformation probing and molecular dynamics were applied to the H⎯(RLDL)4–OH peptide monomer as well as for the most stable structures of dimer, tetramer, tetramer ”the leaf,” and octamer complexes. During these investigations the corresponding MD trajectories were calculated, each of 10 ns and comparative data analy sis was performed. It was shown that each peptide complex structure, except for “the leaf” complex, retain the βsheet conformation as a prevailing one (Fig. 1). It was also noticed that the H⎯(RLDL)4–OH dimer reveals much more stable βsheet conformation in explicit water (Fig. 2) as compared to the same type of structure of the H⎯(RADA)4–OH peptide [6]. This suggests the formation of antiparallel H⎯(RLDL)4– OH βsheet dimers as the stage, which may be recog nized as an initial step of the molecule selfassembling in water (Fig. 3, A and B). That conclusion is fully sup L, aa 14

ported by the results of comparative analysis of the Gibbs free energies, which were calculated for the peptide structures (table). In order to clarify the possible method of protofila ment formation, we studied two forms of the H⎯(RLDL)4–OH tetramer complex. The most stable tetramer structure is composed of two dimers with the contact area, represented explicitly by the hydropho bic side chains of leucine residues (Fig. 3, C). The peculiarity of this structure is determined by an obvi ous presence of fractal surfaces [10]. Therefore, an interaction of two H–(RLDL)4–OH stable tetramers with their planar regions results in formation of a sta ble octamer as the next level fractal structure. The octamer structure reproduces the contact surface topology both in shape and quantity, equal to the pre cursor tetramer structures (Fig. 3, F). This allowed us to assume that the fractal principle [10], indeed, is RMSF, Å 8

4

Monomer Dimer Tetramer Octamer

7 12

2

6

3

10

5

8

4 3

1

6

2

4

1 2

0

1

2

3

4

5

6

7

8

9

10 t, ns

0

L2

R1

L4 D3

L6 R5

L8 D7

R9

L10 L12 L14 L16 D11 R13 D15

Fig. 1. Maximal length (L) of βsheet structures, i.e., the number of nonintercepted amino acid residues in the H⎯(RLDL)4–OH peptide that are characterized by βconformation along the experimental MD trajectory of 10 ns at an integrating step of 0.2 ps. Designations: 1— dimer, 2—tetramer, 3—tetramer“the leaf”, 4—octamer.

Fig. 2. The values of relative mean square fluctuation (RMSF, Å) for the peptide backbone atoms during MD experiments. Initially, H⎯(RLDL)4–OH peptide com plexes contain all the molecules in the βsheet conforma tion. The abscissas correspond to the group of atoms related to the amino acid residue at the given position.

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2 3 A

B

C

E

F

1

D

Fig. 3. Formation of protofilament structures. Numerals and arrows indicate the side substituents in 1 leucine residues, 2 arginine residues, and 3 aspartate residues. Letters designate H⎯(RLDL)4–OH peptide structures in βconformation: A—monomer; B— dimer; C—stable tetramer (protofilament); D—tetramer“the leaf; E—structural involution of tetramer“the leaf; and F— octamer (protofilament). The ways of the peptide selfassembling into the protofilaments are characterized by the structural paths A–B–C–F, A–B–D–E–C–F, and A–B–D–F.

employed for selfassembling of H–(RLDL)4–OH peptide molecules in water. Finally, we revealed the protofilament structure as a stable tetramer complex (Fig. 3, C). The molecular dynamics of the tetramer“the leaf” complex, which is an alternative structure formed by antiparallel peptide dimers stacked with their planar regions (Fig. 3, D), has revealed the involution of the spatial structure of the complex. This finding was used to identify the spe cific way by which the molecules of the H⎯(RLDL)4– OH peptide undergo their reorganization and mutual relocation (Fig. 3, E) in water. It can be concluded that the formation of “ribbonshaped” complexes, such as the H–(RLDL)4–OH tetramer“the leaf” structure, should be regarded as an alternative way of the protofilament structure formation, because during the change in the tetramer spatial structure the hydropho bic leucine residues of two dimers approach each other in front (Fig. 3, E). This path of spatial arrangement of hydrophobic leucine residues corresponds to the spa tial organization of antiparallel dimers found in the most stable form of the H⎯(RLDL)4–OH peptide tet ramer (Fig. 3, C). It is worth mentioning that the most active shifts are observed during the rearrangement of the tet ramer“the leaf” structure. The movements are fea tured by the antiparallel peptide dimers in the complex rather than by single H⎯(RLDL)4–OH molecules. This fact again confirms the special role that has been

proposed for the H⎯(RLDL)4–OH dimer structure in the selfassembling process. In addition, one should not exclude the possibility that two ribbonlike struc tures, such as tetramer“the leaf” complexes, may directly contact each other through the hydrophobic side chains of leucine residues. Practically the above mentioned interactions will result in the formation of a typical H⎯(RLDL)4–OH peptide tetramer, that is a protofilament structure capable of taking part in fur ther filament assembling in accordance with the frac tality principle. Apparently, the selforganization of H ⎯ (RLDL)4–OH peptide structures during the protofilament formation could be achieved by three ways depicted in Fig. 3. The first way presents the interaction between hydrophobic leucine residues in two antiparallel peptide dimers. The next way is imple mented by forming the “ribbonshaped” structure of the H⎯(RLDL)4–OH peptide tetramer“the leaf” complex (Fig. 3, D), which consecutively undergoes reorganization of the peptide dimers imposed within the complex structure (Fig. 3, E). The third way char acterizes the interaction of a pair of tetramer“the leaf” complexes with each other through the hydro phobic surfaces formed by side chains of leucine resi dues. To conclude, the characteristic topology and conformational stability of H⎯(RLDL)4–OH pep tides complexes were found in this work for dimer, tet ramer, and octamer structures. The spatial motility of

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βsheet structures clearly marks the main stages of peptide molecule selforganization and ensures the possibility of assembling the filament nanoparticles in water. ACKNOWLEDGMENTS This work was supported by the Ministry of Educa tion and Science of the Russian Federation (project nos. 02.740.11.5224, 14.740.11.0170, and 16.512.11.2066) in scope of the Federal programs “Scientific and Education Potential of the Innovating Russia for the Years 2009–2013,” “Research and Development Aimed at Scientific and Technological Complex Development of Russia for the Years 2007– 2012,” and the program of the Russian Academy of Sciences “Molecular and Cellular Biology”. REFERENCES 1. Haass, C. and Selkoe, D.J., Nat. Rev. Mol. Cell. Biol., 2007, vol. 8, pp. 101–112.

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2. Zhang, S., Nat. Biotechnol., 2003, vol. 21, pp. 1171– 1178. 3. Danilkovich, A.V., Lipkin, V.M., and Udovichenko, I.P., Rus. J. Bioorg. Chem., 2011, vol. 37, no. 6, pp. 708–712. 4. Danilkovich, A.V., Tikhonov, D.A., Sobolev, E.V., et al., Mat. Biol. Bioinform., 2011, vol. 6, no. 1, pp. 53– 62. 5. Case, D.A., Darden, T.A., Cheatham, T.E. III, et al., AMBER 11, San Francisco: Univ. of California, 2010. 6. Danilkovich, A.V., Sobolev, E.V., Tikhonov, D.A., et al., Mat. Biol. Bioinform., 2011, vol. 6, no. 1, pp. 92– 101. 7. Jorgensen, W.L., Chandrasekhar, J., Madura, J., and Klein, M.L., J. Chem. Phys., 1983, vol. 79, pp. 926– 935. 8. Essman, U., Perera, L., Berkowitz, M.L., et al., J. Chem. Phys., 1995, vol. 19, pp. 8577–8593. 9. Kabsch, W. and Sander, C., Biopolymers, 1983, vol. 22, pp. 2577–2637. 10. Wadhawan, V.K., in Smart Structures: Blurring the Dis tinction between the Living and the NonLiving, New York: Oxford Univ. Press, 2007.

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