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[en] Antifreeze proteins (AFPs) are synthesized by various cold-adapted organisms to enable them to survive in subzero environment. The unique role of AFPs recently attracted enormous interest to develop them as commercial products. In this work, we have studied the antifreeze activity of short helical protein fragments (peptides) instead of the entire antifreeze protein of Antarctic yeast Glaciozyma antarctica. Several short peptide segments were designed according to amino acid sequence of helical region of AFP-1 G.antarctica, which are assumed to be involved in its antifreeze activity. We have demonstrated that short peptide segments derived from yeast AFP possess antifreeze activity and result in modification of the ice crystals growth rates and habits. This strategy has enabled the preparation of short AFP with high antifreeze activity in large amount of quantities at a low cost further opens the chance of developing the commercial potentials of AFPs.(author)
[en] Summary: A cDNA encoding antifreeze protein (AFP84c) was cloned by RT-PCR from the larva of the yellow mealworm Tenebrio molitor. The coding fragment of 252 bp encodes a protein of 84 amino acid residues and was fused to the expression vectors pMAL-c2X and pMAL-p2X. The expression plasmids pMAL-c2X-afp84c and pMAL-p2X-afp84c were constructed and transformed into Escherischia coli strains TBI, respectively. Strategy of optimization of induction conditions were used for expression of the highly disulfide-bonded beta-helix-contained protein with the activity of antifreeze in pMALTM expression system. The target fusion protein was released from the cytoplasm and periplasm by sonication and cold osmotic shock procedure respectively. Recombinant AFP84c was purified by amylose affinity column. The purified target protein displayed a single band in SDS-PAGE. Expressed AFP84c exhibits to increase low temperature resistance of bacteria. (author)
[en] Calcium nitrate in mortars and concrete is used as a multifunctional additive: as set accelerator, plasticizer, long term strength enhancer and as antifreeze admixture. Used binding material and the amount of calcium nitrate, affect the characteristics of the concrete mixture and strength of hardened concrete. The setting time of the initial and the final binding at different temperatures of hardening (+ 20 °C and + 5 °C) of the pastes made of different cements (Portland cement CEM I 42.5 R and Portland limestone cement CEM II/A-LL 42.5 R) and various amounts of calcium nitrate from 1 % until 3 % were investigated. The effect of calcium nitrate on technological characteristics of concrete mixture (the consistency of the mixture, the density, and the amount of air in the mixture), on early concrete strength after 2 and 7 days, as well as on standard concrete strength after 28 days at different temperatures (at + 20 °C and + 5 °C) were analysed. (paper)
[en] Certain organisms survive low temperatures using a range of physiological changes including the production of antifreeze proteins (AFPs), which have evolved to adsorb to ice crystals. Several of these proteins have been purified and shown to also inhibit the crystallization of clathrate hydrates. They have been found to be effective against structure II (sII) hydrates formed from the liquid tetrahydrofuran, sI and sII gas hydrates formed from single gases, as well as sII natural gas hydrates using a mixture of three gases, as assessed using a variety of instrumentation including stirred reactors, differential scanning calorimetry, nuclear magnetic resonance, Raman spectroscopy, and X-ray powder diffraction. For the most part, AFPs are equal to or more effective than the commercial kinetic hydrate inhibitor (KHI) polyvinylpyrolidone, even under field conditions where saline and liquid hydrocarbons are present. Enclathrated gas analysis has revealed that the adsorption of AFPs to the hydrate surface is distinct from tested commercial KHIs and results in properties that should make these proteins more valuable in some field applications. Efforts to overcome the difficulties of recombinant protein production are ongoing, but in silico models of AFP adsorption to hydrates may offer the opportunity to design commercial KHIs for hydrocarbon recovery and transport with all the attributes of these AFP 'green inhibitors', including their benefits for human and environmental safety. (author)
[en] Highlights: • Antifreeze activity is found to be correlated with a change in hydrogen bond dynamics. • The ‘hydration funnel’, i.e. a retardation of hydration bond dynamics toward the ice binding site is found to be a general phenomena for antifreeze proteins. • Cosolutes, such as sodium citrate influence the hydrogen bond dynamics and increase the antifreeze activity of the protein. • Using THz absorption spectroscopy we can state that sodium citrate has a local, direct interaction with the protein. • Whereas changes in the collective hydrogen bond dynamics are a necessary condition for antifreeze activity, the local mechanism determines the size of the antifreeze activity. We combine Terahertz absorption spectroscopy (THz) and molecular dynamics (MD) simulations to investigate the underlying molecular mechanism for the antifreeze activity of one class of antifreeze protein, antifreeze protein type III (AFP-III) with a focus on the collective water hydrogen bond dynamics near the protein. After summarizing our previous work on AFPs, we present a new investigation of the effects of cosolutes on protein antifreeze activity by adding sodium citrate to the protein solution of AFP-III. Our results reveal that for AFP-III, unlike some other AFPs, the addition of the osmolyte sodium citrate does not affect the hydrogen bond dynamics at the protein surface significantly, as indicated by concentration dependent THz measurements. The present data, in combination with our previous THz measurements and molecular simulations, confirm that while long-range solvent perturbation is a necessary condition for the antifreeze activity of AFP-III, the local binding affinity determines the size of the hysteresis.
[en] Two trapping systems were compared in a study in Guatemala during the wet season, May through Dec 2001. Trap/lure combinations consisting of green or yellow-based plastic McPhail-like traps baited with a synthetic 2-component lure (putrescine and ammonium acetate) and 300 mL of propylene glycol antifreeze as a preservative were compared to the traditional glass McPhail baited with torula yeast/borax and 300 mL of water. Both systems captured several key Anastrepha species including Anastrepha ludens Loew, A. obliqua, Macquart, A. serpentina Weidemann, A. striata Schiner, A. distincta Greene, A. fraterculus Weidemann as well as Ceratitis capitata Weidemann. Additionally, 13 other Anastrepha spp. were captured with the synthetic lure. The plastic traps captured more key flies than the McPhail trap except for A. distincta where there were no significant differences between the yellow-based plastic trap and the McPhail trap and no significant differences between any trap and lure for trapping A. fraterculus. The synthetic lure lasted 10 weeks. The sex ratio was female-biased for almost all captured key species in both systems. Moreover, there were significant numbers of captured nontarget insects in all traps; however, the captured flies in those traps with the synthetic lure were not adversely affected by these insects. Propylene glycol-based antifreeze was a superior preservative when compared to borax/water. (author)
[es]En Guatemala, se compararon dos sistemas de trampeo durante la epoca lluviosa de Mayo a Deciembre, 2001. Combinaciones de trampa/atrayente que consistieron de trampas de plastico con bases verdes o amarillos y con atrayentes sinteticos (acetate de amoniaco y putrecina) fueron comparadas con el sistema de trampeo tradicional McPhail de vidrio cebada con torula y borax en agua. Los dos sistemas capturaron moscas del genero Anastrepha incluyendo Anastrepha ludens Loew, A. obliqua, Macquart, A. serpentina Weidemann, A. striata Schiner, A. distincta Greene, A. fraterculus Weidemann y Ceratitis capitata Weidemann. Ademas, se capturaron 13 especias adicionales de Anastrepha asi como Toxotrypana curvicauda Gerstaecker con el cebo sintetico. El cebo sintetico fue efectivo por diez semanas sin recebar. Las trampas de plastico capturaron mas moscas del genero Anastrepha que la trampa de cristal McPhail. Las excepciones fueron A. distincta en donde no hubo differencias el la captura con la trampa de plastico con base amarillo y la trampa McPhail de cristal asi como A. fraterculus en donde no hubo differencias comparando la captura de moscas con ambos sistemas. La proporcion sexual de las moscas capturadas con los dos sistemas fue al favor de las hembras. La captura de otros tipos de insectos fue significantemente elevado, sin embargo, las moscas capturadas con los cebos sinteticos no fueron afectados adversamente por estos insectos. El 10% del anticongelante, glicol propilico, fue superior al borax/agua como conservador de las moscas capturardas. (author)
[en] The dynamical properties of solvation water of hyperactive antifreeze protein from Choristoneura fumiferana (CfAFP) are analyzed and discussed in context of its antifreeze activity. The protein comprises of three well-defined planes and one of them binds to the surface of ice. The dynamical properties of solvation water around each of these planes were analyzed separately; the results are compared with the dynamical properties of solvation water of ice around its two crystallographic planes: basal and prism. Three main conclusions are inferred from our investigations. The first one is that the solvation shell of CfAFP does not seem to be particularly far-ranged, at least not beyond what is usually observed for proteins that do not interact with ice. Therefore, it does not appear to us that the antifreeze activity is enhanced by a long-ranged retardation of water mobility. Also the correlation between the collective mobility of water and the collective mobility of protein atoms highly resembles the one measured for the protein that does not interact with ice. Our second conclusion is that the dynamical properties of solvation water of CfAFP are non-uniform. The dynamics of solvation water of ice-binding plane is, in some respects, different from the dynamics of solvation water of the two remaining planes. The feature that distinguishes the dynamics of solvation water of the three planes is the activation energy of diffusion process. The third conclusion is that—from the three analyzed solvation shells of CfAFP—the dynamical properties of solvation water of the ice-binding plane resemble the most the properties of solvation water of ice; note, however, that these properties still clearly differ from the dynamic properties of solvation water of ice
[en] Solidification of liquid occupying material’s pores is one of the main reasons of its skeleton deterioration. So far, it has been usually assumed that confined water undergoes phase transitions more rapidly than temperature changes, which indicates that the kinetic effects are negligible. However, this assumption is often infringed. The main object of this paper is to analyse the crystallization of commonly applied antifreeze, i.e. ethylene as well as propylene glycol, in water solutions contained in voids of mesoporous silica gel with average pore size equal to 11 nm. In case of both solutions, two concentrations are analysed, 5% and 10%. Additionally, the analysis is conducted also for deionized water. The experimental research has been conducted by means of differential scanning calorimetry with multiple cooling rate program applied. The activation energy is estimated according to the differential Friedman method. In order to pick the most appropriate kinetic model to analysed phenomenon, two approaches are applied: the one introduced by Málek as well as the one described by Perez-Maqueda et al., the so-called model-fitting method. The former indicates the Šesták–Berggren model, whereas the latter points to nth reaction order model. Both equations demonstrate high accordance with the experimental data. The Šesták–Berggren equation is an empirical formula, which does not provide any explanation about antifreeze solidification. However, the nth reaction order model belongs to the group of geometrical extension/contraction models.
[en] The dynamics of threonine side chains of the Tenebrio molitor antifreeze protein (TmAFP) were investigated using natural abundance 13C NMR. In TmAFP, the array of threonine residues on one face of the protein is responsible for conferring its ability to bind crystalline ice and inhibit its growth. Heteronuclear longitudinal and transverse relaxation rates and the 1H-13C NOE were determined in this study. The CαH relaxation measurements were compared to the previously measured 15N backbone parameters and these are found to be in agreement. For the analysis of the threonine side chain motions, the model of restricted rotational diffusion about the χ1 dihedral angle was employed [London and Avitabile (1978) J. Am. Chem. Soc., 100, 7159-7165]. We demonstrate that the motion experienced by the ice binding threonine side chains is highly restricted, with an approximate upper limit of less than ±25 deg