19F核磁共振标签用于肽基脯氨酸构象分析。

IF 3.1 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY
George S M Hanson, Faidra Batsaki, Teagan L Myerscough, Kristin Piché, Ariel Louwrier, Christopher R Coxon
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引用次数: 0

摘要

脯氨酸顺式/反式异构在蛋白质内部无序区域的短线性相互作用基序中,在蛋白质折叠和介导蛋白质相互作用中起重要作用。顺式和反式脯氨酸键之间的缓慢交换速率在氟化肽的19F NMR分析中提供了不同的信号,允许简单地定量每个人口。然而,氟并非天然存在于蛋白质中,但可以通过化学标签引入。在这项研究中,我们评估了一系列氟化半胱氨酸反应性19F核磁共振标签,以评估它们与短的线性脯氨酸肽反应的能力,并准确报告平衡的顺式/反式脯氨酸群体。几种氟化亲电标签,包括硝基苯、磺酰基嘧啶和丙烯酰胺,被发现对模型肽Ac-LPAAC中的%cis-Pro进行化学选择性反应,并可靠地报告。其他19F核磁共振标签被发现是局部脯氨酸构象的差报告者。虽然五氟吡啶是非化学选择性的,但在Ac-LPAAX (X = Cys, Tyr, Lys)多肽中通过半胱氨酸或酪氨酸偶联时,它仍然可靠地报道了%顺式pro。3,4-二氟硝基苯与α-突触核蛋白半胱氨酸突变体反应时产生一对区域异构体标记产物,但与蛋白质标记相容。这些工具可能对探测蛋白质中的顺式/反式亲群体有价值。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
19F NMR-tags for peptidyl prolyl conformation analysis.

Proline cis/trans isomerism plays an important role in protein folding and mediating protein-protein interactions in short linear interacting motifs within intrinsically disordered protein regions. The slow exchange rate between cis and trans prolyl bonds provides distinct signals in 19F NMR analysis of fluorinated peptides, allowing for simple quantification of each population. However, fluorine is not naturally found in proteins but can be introduced using chemical tags. In this study, we evaluate a range of fluorinated cysteine-reactive 19F NMR tags to assess their ability to react with short, linear proline-containing peptides and accurately report on the equilibrium cis/trans-Pro populations. Several fluorinated electrophilic tags, including nitrobenzenes, sulfonylpyrimidines, and acrylamides, were found to react chemoselectively and reliably report on the %cis-Pro in the model peptide Ac-LPAAC. Other 19F NMR tags were found to be poor reporters of local proline conformation. Although pentafluoropyridine was non-chemoselective, it still reliably reported on %cis-Pro when conjugated via cysteine or tyrosine in Ac-LPAAX (X = Cys, Tyr, Lys) peptides. 3,4-Difluoronitrobenzene was found to be compatible with protein tagging, albeit it had modest reactivity and afforded a pair of regioisimeric tagging-products when reacted with a cysteine mutant of α-synuclein. These tools may be valuable for probing cis/trans-Pro populations in proteins.

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来源期刊
CiteScore
6.10
自引率
0.00%
发文量
128
审稿时长
10 weeks
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