Abstract 1452: Modeling the binding of ω-conotoxin and other toxins to the N-type voltage-gated calcium channel
Abstract 1452: Modeling the binding of ω-conotoxin and other toxins to the N-type voltage-gated calcium channel
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摘要 1452：模拟 β-芋螺毒素和其他毒素与 N 型电压门控钙通道的结合
DOI：
10.1016/j.jbc.2023.104024
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发表时间：
2023
影响因子：
4.8
通讯作者：
Sikora, Arthur
中科院分区：
文献类型：
--
作者：
Sha, Serena;Guillen-Tapia, Ashley;Welch, Sophie;Lavin, Emily Schmitt;Sikora, Arthur
嗜热红球藻血红素-一氧化氮和/或氧结合结构域（Cs H-NOX）是一种嗜热血红素蛋白，其结合小气体分子，包括氧气、一氧化氮和一氧化碳。在这里，非典型氨基酸（ncAA）被用来探测蛋白质中的局部溶剂化环境或修饰蛋白质的功能。具体地，使用琥珀密码子抑制方法，将ncAA振动报告基因4-氰基-L-苯丙氨酸（pCNPhe）在多个位点特异性地掺入蛋白质中，包括蛋白质中的表面和埋藏位点。然后利用温度依赖性IR光谱来测量pCNPhe的腈对称伸缩频率的温度依赖性，其与所产生的蛋白质构建体的X射线晶体结构结合的局部溶剂化环境相关。另外，酪氨酸类似物在位点140处单独掺入，位点140是天然蛋白质结构中的酪氨酸。Y140参与与血红素铁结合的氧的氢键相互作用。因此，预测酚氢的pKa的调节会影响这种相互作用的强度，从而影响蛋白质与氧结合的亲和力。将展示用ncAA探测局部蛋白质环境和调节氧亲和力的结果。104023，https：//doi。org/10.1016/j.jbc. 2023.104023
Caldanaerobacter subterraneus heme-nitric oxide and/or oxygen binding domain (Cs H-NOX) is a thermophilic heme protein that binds to small gas molecules including oxygen, nitric oxide, and carbon monoxide. Here, non-canonical amino acids (ncAAs) were utilized to either probe the local solvation environments in the protein or to modify the functionality of the protein. Specifically, the ncAA vibrational reporter 4-cyano-L-phenylalanine (pCNPhe) was site-specifically incorporated into the protein at a number sites using the Amber codon suppression methodology, including surface and buried sites in the protein. Temperature-dependent IR spectroscopy was then utilized to measure the temperature dependence of the nitrile symmetric stretching frequency of pCNPhe which was correlated to local solvation environment in conjunction with X-ray crystal structures of the protein constructs generated. Additionally, tyrosine analogs were individually incorporated at site 140 which is a tyrosine in the native protein structure. Y140 is involved in a hydrogen bonding interaction with oxygen bound to the heme iron. Thus modulation of the pKa of the phenolic hydrogen is predicted to impact the strength of this interaction and thus the affinity of the protein to oxygen binding. Results from probing local protein environments with ncAAs and modulating oxygen affinity will be presented.104023, https://doi. org/10.1016/j. jbc. 2023.104023