Mutational analysis of protein solubility enhancement using short peptide tags
Mutational analysis of protein solubility enhancement using short peptide tags
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DOI：
10.1002/bip.20596
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发表时间：
2007-01-01
期刊：
影响因子：
2.9
通讯作者：
Kuroda, Yutaka
中科院分区：
文献类型：
--
作者：
Kato, Atsushi;Maki, Kosuke;Kuroda, Yutaka
蛋白质聚集是一种普遍现象。制备高度浓缩的蛋白质样品通常是生物物理测量所需的，通常涉及对提高蛋白质溶解度的溶剂条件进行耗时和繁琐的测试。在这里，在系统的分析中，我们已经确定了在我们的低溶解度模型蛋白BPTI-22(一种含有22个丙氨酸的BPTI变体)的N或C末端添加包含1、3和5个赖氨酸或精氨酸(或6个精氨酸)的SEP标签(溶解性增强肽标签)时，溶解度的增加。正如预期的那样，BPTI-22的溶解度与SEP-Tag中包含的带电残基的数量直接相关，而不改变蛋白质的活性或结构。最大的溶解度增加是在C末端添加5个赖氨酸(BPTI-22-C5K)、5个和6个精氨酸残基(BPTI-22-C5R和BPTI-22-C6R)所产生的4.2倍、4.8倍和6.2倍。增加标记的BPTI-22的溶解度可以产生更高质量的核磁共振谱(关于信噪比和线形的异质单量子相关HSQC谱)，比未标记的BPTI-22的时间短得多。此外，通过HSQC光谱观察到，标记的样品在10天以上仍然是可溶的。我们相信，基于赖氨酸和精氨酸的SEP标签可能会为提高蛋白质的溶解度提供一种有效和通用的方法。(C)2006年威利期刊公司。
Protein aggregation is a common phenomenon. The preparation of highly concentrated protein samples, typically required for biophysical measurements, often involves a time consuming and tedious testing of solvent conditions for improving protein solubility. Here, in a systematic analysis, I we have determined the increase in solubility upon the addition of SEP-tags (solubility enhancement peptide tags) containing, one, three, and five lysines or arginines (or six arginines) to either the N or C terminus of our low solubility model protein, bovine pancreatic trypsin inhibitor variant, BPTI-22 (a BPTI variant containing 22 alanines). As anticipated, the BPTI-22 solubility increased in direct relation to the number of charged residues contained in the SEP-tag, and without altering either the activity or the structure of the protein. The largest solubility increases were of 4.2-, 4.8-, and 6.2-folds produced by the addition, at the C terminus, of five lysine (BPTI-22-C5K), five and six arginine residues (BPTI-22-C5R and BPTI-22-C6R), respectively. ne increased solubility of the tagged BPTI-22 yielded higher quality NMR spectra (hetero single quantum correlation HSQC spectra, with respect of the signal-to-noise and line shapes) in, a much shorter time than for the untagged BPTI-22. Furthermore, tagged samples remained soluble for over ten days, as observed by their HSQC spectra. We believe that lysine- and arginine-based SEP-tags may provide an effective and versatile method for enhancing protein solubility. (c) 2006 Wiley Periodicals, Inc.