High-level expression of uniformly 15N-labeled hen lysozyme in Pichia pastoris and identification of the site in hen lysozyme where phosphate ion binds using NMR measurements
High-level expression of uniformly 15N-labeled hen lysozyme in Pichia pastoris and identification of the site in hen lysozyme where phosphate ion binds using NMR measurements
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DOI：
10.1016/s0014-5793(99)00332-4
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发表时间：
1999-04-01
期刊：
影响因子：
3.5
通讯作者：
Imoto, T
中科院分区：
文献类型：
--
作者：
Mine, S;Ueda, T;Imoto, T
已知 Asn 到 Asp 的非酶促脱酰胺作用发生在蛋白质和肽中，并通过磷酸盐缓冲液加速 [Tyler-Cross, R. 和 Schirch, V. (1991) J. Biol.化学。 25、22549-22556]。我们试图通过 N-15 标记的溶菌酶的 H-1-N-15 HSQC 测量来识别溶菌酶中磷酸根离子结合的位点，这是使用巴斯德毕赤酵母成功获得的。结果，我们发现磷酸根离子优先与母鸡溶菌酶中的Asn-103结合。这里提出的方法可能有助于识别蛋白质与低分子量物质的结合位点。 (C) 1999 年欧洲生化学会联合会。
The non-enzymatic deamidation of Asn to Asp is known to occur in proteins and peptides and is accelerated by phosphate buffer [Tyler-Cross, R. and Schirch, V. (1991) J. Biol. Chem. 25, 22549-22556]. We attempted to identify the site in lysozyme where a phosphate ion binds by means of H-1-N-15 HSQC measurements of N-15-labeled lysozyme, which was successfully obtained using Pichia pastoris. As a result, we found that the phosphate ion was preferentially bound to Asn-103 in hen lysozyme, The method presented here may be useful for identifying the binding site of a protein with low molecular weight substances. (C) 1999 Federation of European Biochemical Societies.