Protein Crystallization by Forced Flow through Glass Capillaries: Enhanced Lysozyme Crystal Growth
Protein Crystallization by Forced Flow through Glass Capillaries: Enhanced Lysozyme Crystal Growth
复制标题
通过玻璃毛细管强制流动进行蛋白质结晶：增强溶菌酶晶体生长
DOI：
10.1021/cg900492j
复制
发表时间：
2010
影响因子：
3.8
通讯作者：
Roberts M
中科院分区：
文献类型：
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作者：
Roberts M
蛋白质晶体在静止模式下的生长取决于封闭容器中蛋白质的供应，其中晶体生长到受与晶体接触的溶液中蛋白质质量限制的尺寸。在这里，我们表明，通过诱导溶液流到一个较低的温度环境中，蛋白质晶体生长在毛细管中的速度更快，更大的比在密封的毛细管中的静态条件下获得的。在毛细管流动下，晶体尺寸分布也更规则。在27小时内，溶菌酶晶体在流动条件下生长至0.37 mm，在静止条件下生长至0.16 mm。相对于密封批结晶的晶体质量的75倍浓度，达到了在一个等效的毛细管空间内使用这种流动技术与更大体积的蛋白质溶液。我们利用这一特点开发了一种循环系统，以最大限度地提高通过毛细管流动获得的晶体的产量。通过每个毛细管再循环蛋白质溶液40小时，使晶体产率增加到总蛋白质的80%，接近于密封毛细管分批控制结晶的产率。这种方法为生长用于结构测定的大蛋白质晶体和大规模制备蛋白质治疗剂提供了希望。
The growth of protein crystals in quiescent mode depends on the supply of protein in a closed vessel, where crystals grow to a size limited by the mass of protein in solution in contact with the crystal. Here, we show that by inducing solution flow into a lower temperature environment, protein crystals grow in capillaries faster and larger than those obtained under quiescent conditions in sealed capillaries. The crystal size distribution is also more regular under capillary flow. In 27 h, lysozyme crystals grow up to 0.37 mm under flow conditions and 0.16 mm under quiescent conditions. A 75-fold concentration of crystal mass relative to a sealed batch crystallization was attained within an equivalent capillary space using this flow technique with a greater volume of protein solution. We have exploited this feature to develop a circulatory system to maximize the yield of crystals obtained by capillary flow. Recirculating the protein solution through each capillary for 40 h increases the crystal yield to 80% of total protein, close to that of a sealed capillary batch control crystallization. This method offers promise for growing large protein crystals for use in structure determinations and for the mass preparation of protein therapeutics.