AFPs inhibition

AFP Inhibition of HRP activity when AFP binds to SPCE-surface-immobilised HRP-labelled AFP antibody/ chitosan Decrease in DPV current response for reduction of oxidised thionine (catalyst = H2O2)... 

Antifreeze proteins (AFPs) are ice-binding proteins found in some organisms (such as fish, insects, plants and soil bacteria) that live at the temperature of their surroundings and encounter freezing conditions. AFPs help organisms to survive below 0°C by inhibiting ice growth. AFPs are structurally diverse, each is radically different from the others in its primary,... 

Freeze-tolerant insects and plants have been found to contain substantial concentrations of AFP s, a phenomenon that may appear paradoxical. However, THPs present in the extracellular fluids may contribute importantly to freeze-tolerance by inhibiting recrystallization, thus keeping the ice crystals that do form in the extracellular space small enough to prevent... 

Fig. 1. Standard inhibition curves for the measurement of carcinoembryonic antigen (A), a-fetoprotein (B), and insulin (C) by radioimmunoassays utilizing Z-gel. The total amounts of I in the tubes used for the preparation of the inhibition curve were 90,000 cpm, 79,000 cpm, and 45,000 cpm for carcinoembryonic antigen (CEA), a-fetoprotein (AFP), and insulin, respectively, with a gamma scintillation counter efficiency of 45%.

Fig. 7. Inhibition of binding of mouse anti-a-fetoprotein (AFP) to Microtiter wells coated with AFP by soluble AFP.

Figure 4 Inhibition of ice recrystallization. Samples in 10 pi microcapillaries (740 pm diameter) were frozen and placed at -6°C and examined between crossed polarizing filters. Images were taken prior to, and after overnight incubation, but only the latter images are shown. From left to right samples include sample buffer controls (I, 2), bovine serum albumin, a control protein diluted to 0.2 and 0.02 mg/ ml, respectively (3,4), serial dilutions of 0.2, 0.02 and 0.002 mg/ ml Type I fish antifreeze protein in buffer (5-7), Chryseobacterium sp. cultures (8, 9), and E. coli cultures (10, 11). Note that only the fish AFP and the Chryseobacterium sp. cidtures have crystals too small to be detected at this magnification and the overlying feathery pattern, typical of snap frozen samples, is apparent. Bacterial cultures were at 2 x 10 CFU/ ml. Lines indicate duplicate samples and arrows indicate samples that were diluted.

Of the ten recovered isolates, some showed protein-mediated IR inhibition as well as ice-shaping activities, properties that together are suggestive of AFPs. One of these,... 

Carlo simulation of ice-crystal growth to study the mechanism of inhibition of AFPs on the surface of the THF hydrate and propane hydrate. They found that most of the octahedron surfaces of the THF hydrate were covered with AFP molecules, which could reduce the growth rate of the THF hydrate only allowing plate growth perpendicular to that surface. It is thus necessary to look for other experimental evidences to clarify the common features of AFPs on the inhibition of the clathrate hydrate formation. 

Table 1 summarizes the results obtained for C02-hydrate film formation in AFP-solution experiments. In each column, the upper value is that obtained in the first run, while the lower number in brackets is the average for all subsequent runs with its variation. For AFP-0.01 mg ml-1 samples, we showed the values in brackets are standard deviation> for 8 times repeating experiments. Table 1 shows that rwas estimated as ranging between 60 and 100 min, which is much longer than the periods observed on the pure water droplet. It is concluded that CO2 hydrate formation was clearly inhibited by the presence of AFP molecules, even though the concentration was as small as 10 ppm. 

The inhibition effects of type-III AFP and trehalose, two cryoprotecting materials produced in animals, on type-I CO2 clathrate-hydrates were examined. For comparison with the results of a previous study in which the lateral growth rates of COi-hydrate film were dependent on temperature, pressure and NaCl concentration, the solution droplet was observed in a high pressure vessel filled with CO2. Type-III AFP was found to increase the induction period and to reduce the lateral growth rate of C02-hydrate films. It worked well at low concentrations, indicating that AFP works as a kinetic inhibitor. It was also indicated that AFP would weaken the memory effect of C02-hydrate formation. Trehalose had similar inhibition effects on both the induction period and the lateral growth rate, but it had little apparent concentration-dependence on them. Since trehalose also causes the equilibrium conditions of the CO2 hydrate to shift to lower temperatures, it works not only as a thermodynamic inhibitor but also as a kinetic inhibitor, especially as an anti-agglomerant. 

Although AF(G)Ps and LDHIs are distinct, they both inhibit the growth of crystals. Neither AFGP nor PVP are reported to significantly affect ice nucleation, and similary, we have shown that AFPs and PVP did not affect homogeneous nucleation of THF hydrate. It is not known if these two types of inhibitors can adsorb to other hydrophilic surfaces, however silica is an ubiquitous impurity and common to both these systems. Thus, it is of interest to determine the effects of these inhibitors on heterogeneous nucleation of ice/clathrate hydrate. 

One has to distinguish our studies here from the more popular studies on the adsorption of AFPs or LDHIs on ice or gas hydrate since the former focuses on the effect of AFP/LDHI on heterogeneous nucleation of ice/gas hydrate and the later focuses on the effect on the effects on the ice/hydrate growth. Thus, the studies represented in this study deals with impurity surfaces but not ice/hydrate surface. The authors think that this is an important aspect for the understanding of good ice/hydrate inhibition since the nucleation process is heterogeneous and must involve impurity surfaces. 

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