Tuning crystallization pathways through sequence engineering of biomimetic polymers

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Two-step nucleation pathways in which disordered, amorphous, or dense liquid states precede the appearance of crystalline phases have been reported for a wide range of materials, but the dynamics of such pathways are poorly understood. Moreover, whether these pathways are general features of crystallizing systems or a consequence of system-specific structural details that select for direct versus two-step processes is unknown. Using atomic force microscopy to directly observe crystallization of sequence-defined polymers, we show that crystallization pathways are indeed sequence dependent. When a short hydrophobic region is added to a sequence that directly forms crystalline particles, crystallization instead follows a two-step pathway that begins with the creation of disordered clusters of 10–20 molecules and is characterized by highly non-linear crystallization kinetics in which clusters transform into ordered structures that then enter the growth phase. The results shed new light on non-classical crystallization mechanisms and have implications for the design of self-assembling polymer systems.

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Figure 1: Structures of peptoid molecules and crystals formed on mica.
Figure 2: In situ view of peptoid assembly and resulting morphologies.
Figure 3: Kinetics of crystal formation and bilayer addition.
Figure 4: Comparison of Pepc cluster and crystal formation rates with model predictions.
Figure 5: Proposed peptoid crystallization pathways and energy landscapes.


  1. 1

    Kashchiev, D. Thermodynamically consistent description of the work to form a nucleus of any size. J. Chem. Phys. 118, 1837–1851 (2003).

  2. 2

    Chung, S., Shin, S. H., Bertozzi, C. R. & De Yoreo, J. J. Self-catalyzed growth of S layers via an amorphous-to-crystalline transition limited by folding kinetics. Proc. Natl Acad. Sci. USA 107, 16536–16541 (2010).

  3. 3

    De Yoreo, J. J. et al. Crystallization by particle attachment in synthetic, biogenic, and geologic environments. Science 349, aaa6760 (2015).

  4. 4

    Galkin, O., Chen, K., Nagel, R. L., Hirsch, R. E. & Vekilov, P. G. Liquid–liquid separation in solutions of normal and sickle cell hemoglobin. Proc. Natl Acad. Sci. USA 99, 8479–8483 (2002).

  5. 5

    Nielsen, M. H., Aloni, S. & De Yoreo, J. J. In situ TEM imaging of CaCO3 nucleation reveals coexistence of direct and indirect pathways. Science 345, 1158–1162 (2014).

  6. 6

    Smith, B. J. & Dichtel, W. R. Mechanistic studies of two-dimensional covalent organic frameworks rapidly polymerized from initially homogenous conditions. J. Am. Chem. Soc. 136, 8783–8789 (2014).

  7. 7

    Vekilov, P. G. Two-step mechanism for the nucleation of crystals from solution. J. Cryst. Growth 275, 65–76 (2005).

  8. 8

    Erdemir, D., Lee, A. Y. & Myerson, A. S. Nucleation of crystals from solution: classical and two-step models. Acc. Chem. Res. 42, 621–629 (2009).

  9. 9

    Gebauer, D. & Colfen, H. Prenucleation clusters and non-classical nucleation. Nano Today 6, 564–584 (2011).

  10. 10

    Wallace, A. F. et al. Microscopic evidence for liquid–liquid separation in supersaturated CaCO3 solutions. Science 341, 885–889 (2013).

  11. 11

    Savage, J. & Dinsmore, A. Experimental evidence for two-step nucleation in colloidal crystallization. Phys. Rev. Lett. 102, 198302 (2009).

  12. 12

    Sun, J. & Zuckermann, R. N. Peptoid polymers: a highly designable bioinspired material. ACS Nano 7, 4715–4732 (2013).

  13. 13

    Nam, K. T. et al. Free-floating ultrathin two-dimensional crystals from sequence-specific peptoid polymers. Nat. Mater. 9, 454–460 (2010).

  14. 14

    Mannige, R. V. et al. Peptoid nanosheets exhibit a new secondary-structure motif. Nature 526, 415–420 (2015).

  15. 15

    Chen, C.-L., Zuckermann, R. N. & De Yoreo, J. J. Surface-directed assembly of sequence-defined synthetic polymers into networks of hexagonally-patterned nanoribbons with controllable functionalities. ACS Nano 10, 5314–5320 (2016).

  16. 16

    Kühnle, R. I. & Börner, H. G. Calcium ions to remotely control the reversible switching of secondary and quaternary structures in bioconjugates. Angew. Chem. Int. Ed. 50, 4499–4502 (2011).

  17. 17

    Pashley, R. M. & Quirk, J. P. The effect of cation valency on DLVO and hydration forces between macroscopic sheets of muscovite mica in relation to clay swelling. Colloid Surf. 9, 1–17 (1984).

  18. 18

    Xu, L. & Salmeron, M. An XPS and scanning polarization force microscopy study of the exchange and mobility of surface ions on mica. Langmuir 14, 5841–5844 (1998).

  19. 19

    Friddle, R. W., Noy, A. & De Yoreo, J. J. Interpreting the widespread nonlinear force spectra of intermolecular bonds. Proc. Natl Acad. Sci. USA 109, 13573–13578 (2012).

  20. 20

    Friddle, R. W., Podsiadlo, P., Artyukhin, A. B. & Noy, A. Near-equilibrium chemical force microscopy. J. Phys. Chem. C 112, 4986–4990 (2008).

  21. 21

    De Yoreo, J. J. & Vekilov, P. G. in Biomineralization Vol. 54 Reviews in Mineralogy Geochemistry (eds Dove, P. M., De Yoreo, J. J. & Weiner, S.) 57–93 (Mineralogical Society of America, 2003).

  22. 22

    De Yoreo, J. Crystal nucleation: more than one pathway. Nat. Mater. 12, 284–285 (2013).

  23. 23

    Pouget, E. M. et al. The initial stages of template-controlled CaCO3 formation revealed by Cryo-TEM. Science 323, 1455–1458 (2009).

  24. 24

    Byington, M. C., Safari, M. S., Conrad, J. C. & Vekilov, P. G. Protein conformational flexibility enables the formation of dense liquid clusters: tests using solution shear. J. Phys. Chem. Lett. 7, 2339–2345 (2016).

  25. 25

    Vorontsova, M. A., Chan, H. Y., Lubchenko, V. & Vekilov, P. G. Lack of dependence of the sizes of the mesoscopic protein clusters on electrostatics. Biophys. J. 109, 1959–1968 (2015).

  26. 26

    Gebauer, D., Volkel, A. & Colfen, H. Stable prenucleation calcium carbonate clusters. Science 322, 1819–1822 (2008).

  27. 27

    Addadi, L., Raz, S. & Weiner, S. Taking advantage of disorder: amorphous calcium carbonate and its roles in biomineralization. Adv. Mater. 15, 959–970 (2003).

  28. 28

    Gal, A., Weiner, S. & Addadi, L. A perspective on underlying crystal growth mechanisms in biomineralization: solution mediated growth versus nanosphere particle accretion. CrystEngComm 17, 2606–2615 (2015).

  29. 29

    Du, W., Cruz-Cabeza, A. J., Woutersen, S., Davey, R. J. & Yin, Q. Can the study of self-assembly in solution lead to a good model for the nucleation pathway? The case of tolfenamic acid. Chem. Sci. 6, 3515–3524 (2015).

  30. 30

    Zuckermann, R. N., Kerr, J. M., Kent, S. B. H. & Moos, W. H. Efficient method for the preparation of peptoids [oligo(N-substituted glycines)] by submonomer solid-phase synthesis. J. Am. Chem. Soc. 114, 10646–10647 (1992).

  31. 31

    Friddle, R. W. et al. Single-molecule determination of the face-specific adsorption of Amelogenin’s C-terminus on hydroxyapatite. Angew. Chem. Int. Ed. 50, 7541–7545 (2011).

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Peptoid synthesis, MD simulations, in situ AFM, DFS and TEM characterization and high-speed AFM imaging were supported by the US Department of Energy, Office of Basic Energy Sciences, Biomolecular Materials Program at Pacific Northwest National Laboratory (PNNL) and the Lawrence Livermore National Laboratory. Development of MD potentials and peptoid designs was supported by the Materials Synthesis and Simulation Across Scales Initiative through the LDRD Program at PNNL. PNNL is a multi-program national laboratory operated for Department of Energy by Battelle under Contract No. DE-AC05-76RL01830. Work at the Lawrence Livermore National Laboratory was performed under the auspices of the US Department of Energy under Contract DE-AC52-07NA27344.

Author information

X.M. performed AFM and DLS experiments, data analysis and manuscript writing; S.Z. performed AFM experiments, data analysis and manuscript writing; F.J. performed DFS measurements; C.J.N. performed cryoTEM imaging: Y.Z. performed high-speed AFM imaging; A.P. performed MD simulations; Z.L. performed peptoid synthesis and characterization; M.D.B. performed MD simulations and manuscript writing; C.J.M. designed the simulations; J.P. designed the simulations; A.N. designed the high-speed AFM experiments and performed manuscript writing; C.-L.C. designed the study and performed peptoid design, synthesis and characterization, and manuscript writing; J.J.D.Y. designed the study, developed and applied the mathematical model, and performed data analysis and manuscript writing.

Correspondence to Chun-Long Chen or James J. De Yoreo.

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Ma, X., Zhang, S., Jiao, F. et al. Tuning crystallization pathways through sequence engineering of biomimetic polymers. Nature Mater 16, 767–774 (2017) doi:10.1038/nmat4891

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