Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria, 3086, Australia.
Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria, 3086, Australia.
Protein Expr Purif. 2020 Jul;171:105626. doi: 10.1016/j.pep.2020.105626. Epub 2020 Mar 17.
Splicing factor proline- and glutamine-rich (SFPQ) is an RNA-binding protein, playing significant roles in gene regulation and subnuclear body formation. Our recent serendipitous discovery showed that SFPQ binds zinc directly and forms an infinite polymer that is induced by zinc binding to the protein. The zinc-induced reversible polymerization has led us to exploit this property to develop a rapid purification strategy for SFPQ without the use of affinity tags. In combination with the variation of ionic strength for salting-out of SFPQ, the reversible zinc-induced precipitation of SFPQ reduced the purification time required to obtain pure SFPQ to a single day. The purified protein was subjected to the previously reported crystallization condition. The resulting crystals diffracted to 2.22 Å resolution, confirming the quality of SFPQ purified with this new rapid purification strategy.
拼接因子脯氨酸和谷氨酰胺丰富(SFPQ)是一种 RNA 结合蛋白,在基因调控和亚核小体形成中发挥重要作用。我们最近偶然发现 SFPQ 直接结合锌,并形成一个无限的聚合物,该聚合物是由锌结合到蛋白质上诱导产生的。锌诱导的可逆聚合使我们能够利用这一特性开发一种快速纯化 SFPQ 的策略,而无需使用亲和标签。结合离子强度的变化进行 SFPQ 的盐析,锌诱导的 SFPQ 可逆沉淀减少了获得纯 SFPQ 所需的纯化时间到一天。纯化后的蛋白质进行了先前报道的结晶条件。得到的晶体衍射到 2.22Å 的分辨率,证实了用这种新的快速纯化策略纯化的 SFPQ 的质量。