HIGH PRESSURE CRYOCOOLING AND RADIATION DAMAGE

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得主要资金， 因此可以在其他CRISP条目中表示。列出的机构是 研究中心，而研究中心不一定是研究者所在的机构。 生物样品的辐射损伤是目前大分子X射线晶体学研究的主要限制因素之一。在室温下减少辐射损伤的最佳已知方法之一是将晶体快速冷冻到低温温度，这通常需要寻找渗透性冷冻保护剂。近年来发展起来的一种新的晶体冷冻方法--高压低温冷却法，通过在高压（1000 ~ 2000 atm）下冷冻晶体而避免了使用冷冻保护剂。建议的机制涉及高密度无定形（HDA）冰，在环境压力下的密度为1.17克/厘米3。 由于二次辐射损伤高度依赖于由X射线辐射产生的自由基的扩散，HDA冰可能有效地减少辐射损伤，这是由于与通常在环境压力下在晶体内部形成的低密度无定形冰相比，HDA冰具有高粘度和高密度。 我们将测试高压低温冷却对辐射损伤的影响。在这项研究中，将在几种不同的压力（1 atm ~ 4000 atm）和不同的化学条件下制备高质量的蛋白质晶体。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Radiation damage to biological samples is currently one of the major limiting factors in macromolecular x-ray crystallography. One of the best known method to reduce radiation damage at room temperature is flash freezing crystals to cryogenic temperature, which often requires finding penetrating cryoprotectants. Recently a new crystal freezing method, high pressure cryocooling, was developed, where using of cryoprotectants was avoided by freezing crystals under high pressures (1000 ~ 2000 atm). The mechanism was suggested involving high density amorphous (HDA) ice which has density of 1.17 g/cm3 at ambient pressure. Since the secondary radiation damage highly depends on the diffusion of the free radicals produced by x-ray radiation, HDA ice might be effective to reduce radiation damage due to its high viscosity and density compared to low density amorphous ice that usually forms inside crystal at ambient pressure. We will test the effect of high pressure cryocooling on radiation damage. For the study, high quality protein crystals will be prepared at several different pressures(1 atm ~ 4000 atm) and different chemical conditions.