Research ArticleCHEMICAL BIOLOGY

Ice-nucleating bacteria control the order and dynamics of interfacial water

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Science Advances  22 Apr 2016:
Vol. 2, no. 4, e1501630
DOI: 10.1126/sciadv.1501630
  • Fig. 1 SFG spectra of P. syringae.

    (A to E) SFG spectra of P. syringae bacteria lysate, which contain ice-active inaZ proteins (A) and control substances (B to E) in contact with water at different temperatures. INPs increase the water signal with decreasing temperatures, whereas control substances leave the water signal unchanged. a.u., arbitrary units.

  • Fig. 2 Analysis of temperature-dependent SFG spectra for P. syringae at a water surface.

    (A) Spectra for RT and 5°C along with two fitting components related to more weakly and strongly hydrogen-bonded water within the broad water spectrum. CH and nonresonant fitting components are not included here. (B) Plots of the amplitudes obtained from fits to the SFG spectra from RT to 5°C. The mode related to more strongly hydrogen-bonded water increases strongly, whereas the weakly hydrogen-bonded water mode remains almost unchanged.

  • Fig. 3 MD simulation of the inaZ ice-active site.

    (A) The top view illustrates the ladder-type regions of groups of amino acids on the IN dimer. The side view shows an MD snapshot of the water structure at the IN site. Together, side chains and clathrate water form a template for ice nucleation. Threonine, purple; serine, yellow; alanine, blue; tyrosine, green; glutamic acid, orange; glycine, white. (B) Calculated SFG spectra for regions with different amino acids present. Thr- and Ser-rich areas leave the water signal intensity unchanged, whereas there is a clear trend toward a stronger water signal near glutamic acid– and serine-rich regions. (C) Integrated SFG intensity for the IN site regions at 270 and 300 K. The increased intensity at regions 1 and 6 indicates more ordered water near the perimeter of the IN site.

  • Fig. 4 Energy transfer processes at the water-INP interface.

    (A) Time-resolved difference sum frequency spectrum for the water–P. syringae interface after excitation with a 2470-cm−1 pump pulse near the weakly H-bonded water resonances (dashed line). The signal bleach is very intense in the low-frequency water peak related to strongly H-bonded water. This shows that energy transfer is very rapid and efficient. For clarity, any spectral changes due to thermal effects have been removed. (B) Time-dependent bleach integrated over two spectral regions, 2330 to 2430 cm−1 (strongly H-bonded) and 2480 to 2580 cm−1 (weakly H-bonded). Fits of the data using a coupled differential equation model reveal extremely efficient (80 ± 50 fs) energy transfer between more weakly and more strongly H-bonded water molecules. (C) Time-resolved populations of the more weakly and more strongly H-bonded water molecules extracted from the coupled differential equations. The states become populated from the excitation pulse, energy transfer, and decay to the ground states (not plotted). For the water–P. syringae interface, the more strongly H-bonded state’s population is higher than that of the initially excited peak, which proves extremely efficient energy transfer.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/2/4/e1501630/DC1

    Supplementary Text

    fig. S1. Fitting results for SFG spectra.

    fig. S2. SFG spectra recorded in the amide I region for P. syringae at the air-water interface at RT and 5°C.

    fig. S3. Time traces of surface tension changes recorded while the temperature was changed in the trough for D2O and Snomax at the air-water interface.

    fig. S4. Surface tension changes relative to the tension at 22°C for pure D2O and the Snomax sample for sample temperatures studied with SFG.

    fig. S5. XPS spectra of P. syringae films adsorbed at the water-air interface at 22° and 5°C and transferred to gold-coated silicon chips following the Langmuir-Schaefer (54) method.

    fig. S6. XPS survey spectrum of the Snomax sample.

    fig. S7. Droplet freeze assay for the P. syringae sample used in this study.

    fig. S8. Illustration of the simulation box.

    fig. S9. Calculated SFG intensity Formula for the O–D stretch chromophores within 15 Å of the center of mass of the IN dimer in the z direction.

    fig. S10. Snapshot of the MD simulation near the Thr-rich region (Fig. 3, region 2, main text) of an IN site.

    fig. S11. Time-resolved difference sum frequency spectra for control substances.

    fig. S12. Energy transfer processes at the water-AFP interface.

    table S1. Elemental compositions (atom %) determined by XPS of protein monolayers deposited onto gold-coated silicon chips with the Langmuir-Schaefer technique.

    table S2. Time scales (in femtoseconds) extracted from the coupled differential equation fits (only Formula was allowed to float; the others were fixed).

    References (5568)

  • Supplementary Materials

    This PDF file includes:

    • Bicinonic Acid Assay (BCA)
    • Surface tension experimental details
    • Surface tensions for control samples at RT
    • XPS experimental details
    • fig. S1. Fitting results for SFG spectra.
    • fig. S2. SFG spectra recorded in the amide I region for P. syringae at the air-water
      interface at RT and 5°C.
    • fig. S3. Time traces of surface tension changes recorded while the temperature
      was changed in the trough for D2O and Snomax at the air-water interface.
    • fig. S4. Surface tension changes relative to the tension at 22°C for pure D2O and
      the Snomax sample for sample temperatures studied with SFG.
    • fig. S5. XPS spectra of P. syringae films adsorbed at the water-air interface at 22°
      and 5°C and transferred to gold-coated silicon chips following the Langmuir-Schaefer (54) method.
    • fig. S6. XPS survey spectrum of the Snomax sample.
    • fig. S7. Droplet freeze assay for the P. syringae sample used in this study.
    • fig. S8. Illustration of the simulation box.
    • fig. S9. Calculated SFG intensity | χ xxz2(ω)|2) for the O–D stretch chromophores
      within 15 Å of the center of mass of the IN dimer in the z direction.
    • fig. S10. Snapshot of the MD simulation near the Thr-rich region (Fig. 3, region
      2, main text) of an IN site.
    • fig. S11. Time-resolved difference sum frequency spectra for control substances.
    • fig. S12. Energy transfer processes at the water-AFP interface.
    • table S1. Elemental compositions (atom %) determined by XPS of protein monolayers deposited onto gold-coated silicon chips with the Langmuir-Schaefer technique.
    • table S2. Time scales (in femtoseconds) extracted from the coupled differential equation fits (only down was allowed to float; the others were fixed).
    • References (55–68)

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