We study the properties of water at the surface of an antifreeze protein with femtosecond surface sum frequency generation spectroscopy. We find clear evidence for the presence of ice-like water layers at the ice-binding site of the protein in aqueous solution at temperatures above the freezing point. Decreasing the temperature to the biological working temperature of the protein (0°C to −2°C) increases the amount of ice-like water, while a single point mutation in the ice-binding site is observed to completely disrupt the ice-like character and to eliminate antifreeze activity. Our observations indicate that not the protein itself but ordered ice-like water layers are responsible for the recognition and binding to ice.antifreeze proteins | sum frequency generation | protein hydration

Structure of a bacterial adhesin reveals its role in forming a mixed-species symbiotic community with diatoms on sea ice.

Antibody-based molecular recognition plays a dominant role in the life sciences ranging from applications in diagnostics and molecular imaging to targeted drug delivery and therapy. Here we report a generic approach to introduce protease sensitivity into antibody-based targeting by taking advantage of the intrinsic ability of antibodies to engage in multivalent interactions. Bivalent peptide ligands with dsDNA as a rigid linker were shown to effectively bridge the relatively large distance between the two antigen binding sites within the same antibody, yielding exclusively the cyclic 1 : 1 antibody-ligand complex. Size exclusion chromatography and small angle X-scattering were used to study the types of complexes formed between a model antibody and peptide-dsDNA conjugates displaying 1 or 2 peptide ligands and different linker lengths. Competitive binding assays using fluorescence anisotropy revealed that the interaction between bivalent peptide-dsDNA conjugate and antibody is 500-fold stronger than that of the monovalent peptide, allowing effective blocking of the antigen binding sites in a non-covalent manner. Cleavage of the linker between the peptide epitope and the DNA by matrix metalloprotease 2 disables this strong bivalent interaction and was shown to effectively restore the binding activity of the antibody in an in vitro binding assay. The approach presented here is broadly applicable, because it takes advantage of the Y-shaped multivalent presentation of antigen binding sites common to all antibodies and could be extended to control antibody activity by other input signals.

Poly(vinyl alcohol) (PVA) is a water‐soluble synthetic polymer well‐known to effectively block the recrystallization of ice. The effect of polymer chain architecture on the ice recrystallization inhibition (IRI) by PVA remains unexplored. In this work, the synthesis of PVA molecular bottlebrushes is described via a combination of atom‐transfer radical polymerization and reversible addition‐fragmentation chain‐transfer polymerization. The facile preparation of the PVA bottlebrushes is performed via the selective hydrolysis of the chloroacetate esters of the poly(vinyl chloroacetate) (PVClAc) side chains of a PVClAc precursor bottlebrush. The IRI efficacy of the PVA bottlebrush is quantitatively compared to linear PVA. The results show that even if the PVA chains are densely grafted onto a rigid polymer backbone, the IRI activity of PVA is maintained, demonstrating the flexibility in PVA polymer chain architecture for the design of synthetic PVA‐based ice growth inhibitors.

Articles you may be interested inExplicit-water theory for the salt-specific effects and Hofmeister series in protein solutions J. Chem. Phys. 144, 215101 (2016) Ice binding proteins (IBPs) are produced by various cold-adapted organisms to protect their body tissues against freeze damage. First discovered in Antarctic fish living in shallow waters, IBPs were later found in insects, microorganisms, and plants. Despite great structural diversity, all IBPs adhere to growing ice crystals, which is essential for their extensive repertoire of biological functions. Some IBPs maintain liquid inclusions within ice or inhibit recrystallization of ice, while other types suppress freezing by blocking further ice growth. In contrast, ice nucleating proteins stimulate ice nucleation just below 0 C. Despite huge commercial interest and major scientific breakthroughs, the precise working mechanism of IBPs has not yet been unraveled. In this review, the authors outline the state-of-the-art in experimental and theoretical IBP research and discuss future scientific challenges. The interaction of IBPs with ice, water and ions is examined, focusing in particular on ice growth inhibition mechanisms.

The formation of large ice crystals via recrystallization processes in foods and water-based materials often decreases the quality and structural integrity of the materials. Hence, there is a widespread academic and commercial interest in natural and synthetic ice crystal growth modifiers which inhibit the recrystallization of ice. Well-known natural ice crystal growth modifiers are antifreeze proteins (AFPs), which inhibit ice recrystallization by adsorbing on the surface of ice crystals. Reliable quantification of the ice recrystallization inhibition (IRI) efficiency is a long-sought goal. In this work, we describe a simple method to quantitatively evaluate IRI efficiency, based on automated image analysis using the circle Hough transform (CHT) algorithm. It enables robust and high throughput analysis of natural and synthetic ice recrystallization inhibitors. Here we use the method to evaluate the impact of a single point mutation in the ice-binding site of QAE on its IRI activity. We find that the T18N mutant of QAE has virtually the same effective ice recrystallization inhibitory concentration as the wild-type QAE. This is in contrast to thermal hysteresis activity, evaluated by cryoscopy or sonocrystallization, where the mutation greatly decreases the activity.INTRODUCTION A predictive understanding of (re)crystallization processes would allow to create nanomaterials with novel or enhanced physical and chemical properties. 1 For example, aligned porous materials and other complex structured polymer-inorganic composites have been created by directional freezing methods. [2][3][4] Conversely, ice recrystallization can have a major detrimental impact on the quality and performance of many water-based materials such as foods, biological materials, paints. 5-7 The force driving ice recrystallization is a lowering in the free energy of the system by a reduction in crystal/solution interface energy, due to; isomass, accretion and migration. 8, 9 Isomass recrystallization involves the change in internal structure of ice crystals, and a reduction of crystal defects and surface irregularities. Accretive recrystallization describes the fusion of two neighboring ice crystals. The dominant mechanism at high sucrose concentration is migratory recrystallization (i.e., Ostwald ripening), wherein the mean ice crystal size increases while the number of ice crystals decreases, at a constant volume of ice. The inhibitory effect of antifreeze proteins

We performed time- and polarization-resolved pump-probe and two-dimensional infrared (2D-IR) experiments to study the dynamics of the amide I vibration of a 7 kDa type-III antifreeze protein. In the pump-probe experiments, we used femtosecond mid-infrared pulses to investigate the vibrational relaxation dynamics of the amide mode. The transient spectra show the presence of two spectral components that decay with different lifetimes, indicative of the presence of two distinct amide subbands. The 2D-IR experiments reveal the coupling between the two bands in the form of cross-peaks. On the basis of previous work by Demirdöven et al. ( J. Am. Chem. Soc. 2004 , 126 , 7981 - 7990 ), we assign the observed bands to the two infrared-active modes α(-) and α(+) found in protein β-sheets. The amplitudes of the cross-peak were found to increase with delay time, indicating that the cross-peaks originate from population transfer between the coupled amide oscillators. The time constant of the energy transfer was found to be 6-7 ps.