Relationship of amino acid composition and molecular weight of antifreeze glycopeptides to non-colligative freezing point depression

Many polar fishes synthesize a group of eight glycopeptides that exhibit a non-colligative lowering of the freezing point of water. These glycopeptides range in molecular weight between 2600 and 33700. The largest glycopeptides [1–5] lower the freezing point more than the small ones on a weight basis and contain only two amino acids, alanine and threonine, with the disaccharide galactose-N-acetyl-galactosamine attached to threonine. The smaller glycopeptides, 6, 7, and 8, also lower the freezing point and contain proline, which periodically substitutes for alanine. Glycopeptides with similar antifreeze properties isolated from the saffron cod and the Atlantic tomcod contain an additional amino acid, arginine, which substitutes for threonine in glycopeptide 6. In this study we address the question of whether differences in amino acid composition or molecular weight between large and small glycopeptides are responsible for the reduced freezing point depressing capability of the low molecular weight glycopeptides. The results indicate that the degree of amino acid substitutions that occur in glycopeptides 6–8 do not have a significant effect on the unusual freezing point lowering and that the observed decrease in freezing point depression with smaller glycopeptides can be accounted for on the basis of molecular weight.     

Alterations in membrane transport properties by freezing injury in herbaceous plants:

Leakage of ions from a thawed tissue is a common phenomenon of freezing injury. This leakage is usually assumed to be due to loss of membrane semipermeability or membrane rupture by freezing injury. Freeze injured, yet living, onion (Allium cepa L.) epidermal cells were used to study alterations in cell membranes that result in leakage of ions. In spite of a large efflux of ions, freeze injured cells could be plasmolysed and they remained plasmolysed for several days just like the unfrozen control cells. Injured cells also exhibited protoplasmic streaming. Passive transport of KCl, urea and methyl urea across the cell membranes of injured and control cells was also studied. No difference could be detected for the transport rates of urea and methyl urea between control and injured cells. However, a dramatic increase in the transport rate of KCl was found for the injured cells. Depending upon the extent of initial freezing injury, an increase or a decrease in injury symptoms was found in the post-thaw period. During the progress of freezing injury, 10 days after thawing, a swelling of the protoplasm was seen in the irreversibly injured cells. In spite of this swelling, these cells could be plasmolysed. It appears that the high amount of K⁺ that leaks out into the extracellular water, due to freezing injury, causes protoplasmic swelling by replacing Ca²⁺ in the plasma membrane. We conclude that protoplasmic swelling is a sign of secondary injury. The results presented in this study show that membrane semipermeability is not completely lost and membrane rupture does not occur during the initial stage of freezing injury. In fact, the cells have the ability to repair damage depending upon the degree of injury. Our results show there are specific alterations in membrane semipermeability (e.g., transport of K⁺) which could be repaired completely depending on the degree of injury. These findings suggest that ion leakage due to freezing injury is due to alteration in the membrane proteins and not in the membrane lipids.     

Cryopreservation of marine thraustochytrids (Labyrinthulomycetes)

In this research, the viability of three marine thraustochytrid isolates (fungoid protists) (WSG05, W15 and WH3) were investigated after freezing in liquid nitrogen. Five cryopreservative combinations containing horse serum, glycerol and dimethylsulfide (Me₂SO) were used. The thraustochytrids were assessed directly after removal from liquid nitrogen and cell concentration measured for 10 days post-thawing. Results indicated that a combination of horse serum and Me₂SO were the most effective cryoprotectants for each of the strains tested. Glycerol was only successful in producing growth in one of the strains once thawed.The protocols developed and tested in this study may have further application for cryopreserving other isolates in this class.     

Influence of freezable/non-freezable water and sucrose on the viability of <Emphasis Type="Italic">Theobroma cacao</Emphasis> somatic embryos following desiccation and freezing

Encapsulated cocoa (Theobroma cacao L.) somatic embryos subjected to 0.08–1.25 M sucrose treatments were analyzed for embryo soluble sugar content, non-freezable water content, moisture level after desiccation and viability after desiccation and freezing. Results indicated that the higher the sucrose concentration in the treatment medium, the greater was the extent of sucrose accumulation in the embryos. Sucrose treatment greatly assisted embryo post-desiccation recovery since only 40% of the control embryos survived desiccation, whereas a survival rate of 60–95% was recorded for embryos exposed to 0.5–1.25 M sucrose. The non-freezable water content of the embryos was estimated at between 0.26 and 0.61 g H₂O g⁻¹dw depending on the sucrose treatment, and no obvious relationship could be found between the endogenous sucrose level and the amount of non-freezable water in the embryos. Cocoa somatic embryos could withstand the loss of a fraction of their non-freezable water without losing viability following desiccation. Nevertheless, the complete removal of potentially freezable water was not sufficient for most embryos to survive freezing.     

Antifreeze proteins in the grubby sculpin,Myoxocephalus aenaeus and the tomcod,Microgadus tomcod: comparisons of seasonal cycles

Synopsis Antifreeze protein levels in the plasma of the grubby sculpin, Myoxocephalus aenaeus and the tomcod, Microgadus tomcod of Long Island coastal waters start to increase by November in anticipation of midwinter freezing conditions. Peak levels of antifreeze, as measured by the difference in plasma melting and freezing points, were detected in January for both species. The thermal hysteresis values reached 0.459°C in sculpin and 0.51°C in tomcod. Antifreeze peptides and glycopeptides start to disappear when water temperatures begin to rise and are at insignificant levels by late spring. Aspects of the seasonal cycle and the level of antifreeze activity were compared in three sympatric species (sculpin, tomcod, flounder); in two closely related but ecologically distinct gadids (tomcod, Atlantic cod); and within the genus Myoxocephalus.     

The relationship between freezing tolerance and thermotropic leaf movement in five Rhododendron species

Summary Leaf movement kinetics in five species of Rhododendron were studied in response to leaf temperature, leaf freezing point, and leaf water deficit. There was a gradient in the degree of leaf curling among species in the following order from the greatest curling to the least curling: Rhododendron catawbiense, R. maximum, R. minus, R. macrophyllum, R. ponticum. Those species found to be tolerant of winter conditions had the most intense leaf movements (both curling and angle) while those species with minimal cold tolerance had limited or no leaf movements. Leaf curling occurred at leaf temperatures above the tissue freezing points in all species. Athough leaf angle was influenced by leaf turgor, general tissue desiccation was not the ultimate cause for thermotropic leaf curling in any species tested. Those species with the greatest leaf curling and angle movements had the highest osmotic potential, the lowest water deficit at the turgor loss point, and the lowest symplastic water fraction. These data suggest that there is a trade off in Rhododendron leaf physiology between cold tolerance (due to leaf movements) and water stress tolerance (due to turgor maintenance mechanisms).     

冰冻胁迫下树木管状分子内腔隙和栓塞的形成及其修复

许多树木管状分子细胞的细胞壁在冰冻期间并不随细胞内的水分迁移到细胞外冰晶上而塌陷。此时细胞内产生负压，负压的产生引起腔隙的形成，腔隙又会引起栓塞，导致树木内水分运输受阻。冻融循环可导致腔隙和栓塞的形成，或者冰冻之后，温度急剧回升时树木组织内的冰晶升华所致。在春季树木的根压得到恢复，从而使腔隙和栓塞部分消除，水分运输又得以畅勇。冰冻胁迫对在高纬度和中高纬度的某些地区的木的生长造成很大的危害，管状分子内腔隙和栓塞的形成就是其中之一，也是引起树木生长衰退或死亡的主要原因。本文对腔隙和栓塞的形成的原因，机理及其恢复进行了综述。