Wowk Brian, Fahy Gregory M
21st Century Medicine, Inc., 10844 Edison Court, Rancho Cucamonga, CA 91730, USA.
Cryobiology. 2002 Feb;44(1):14-23. doi: 10.1016/S0011-2240(02)00008-1.
The simple linear polymer polyglycerol (PGL) was found to apparently bind and inhibit the ice nucleating activity of proteins from the ice nucleating bacterium Pseudomonas syringae. PGL of molecular mass 750 Da was added to a solution consisting of 1 ppm freeze-dried P. syringae 31A in water. Differential ice nucleator spectra were determined by measuring the distribution of freezing temperatures in a population of 98 drops of 1 microL volume. The mean freezing temperature was lowered from -6.8 degrees C (control) to -8.0,-9.4,-12.5, and -13.4 degrees C for 0.001, 0.01, 0.1, and 1% w/w PGL concentrations, respectively (SE < 0.2 degrees C). PGL was found to be an ineffective inhibitor of seven defined organic ice nucleating agents, whereas the general ice nucleation inhibitor polyvinyl alcohol (PVA) was found to be effective against five of the seven. The activity of PGL therefore seems to be specific against bacterial ice nucleating protein. PGL alone was an ineffective inhibitor of ice nucleation in small volumes of environmental or laboratory water samples, suggesting that the numerical majority of ice nucleating contaminants in nature may be of nonbacterial origin. However, PGL was more effective than PVA at suppressing initial ice nucleation events in large volumes, suggesting a ubiquitous sparse background of bacterial ice nucleating proteins with high nucleation efficiency. The combination of PGL and PVA was particularly effective for reducing ice formation in solutions used for cryopreservation by vitrification.
研究发现，简单线性聚合物聚甘油（PGL）能明显结合并抑制冰核细菌丁香假单胞菌中蛋白质的冰核活性。将分子量为750 Da的PGL添加到由1 ppm冻干的丁香假单胞菌31A溶于水中形成的溶液中。通过测量98滴1微升体积液滴群体中的冷冻温度分布来确定差异冰核光谱。对于0.001%、0.01%、0.1%和1%（w/w）的PGL浓度，平均冷冻温度分别从-6.8℃（对照）降至-8.0℃、-9.4℃、-12.5℃和-13.4℃（标准误差<0.2℃）。研究发现PGL对七种特定的有机冰核剂是无效抑制剂，而通用冰核抑制剂聚乙烯醇（PVA）对七种中的五种有效。因此，PGL的活性似乎对细菌冰核蛋白具有特异性。单独的PGL对少量环境或实验室水样中的冰核形成是无效抑制剂，这表明自然界中大多数冰核污染物可能是非细菌来源。然而，在抑制大量水样中的初始冰核形成事件方面，PGL比PVA更有效，这表明存在普遍的、具有高成核效率的细菌冰核蛋白稀疏背景。PGL和PVA的组合在减少玻璃化冷冻保存所用溶液中的冰形成方面特别有效。
