DIRECT OBSERVATION OF NITROSYLATED HEME IN MYOGLOBIN AND HEMOGLOBIN BY

直接观察肌红蛋白和血红蛋白中的亚硝化血红素

• 批准号: 6307621
• 负责人: RITA K UPMACIS
• 金额: $ 0.82万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-12-01 至 2000-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6307621
• 关键词: biological products; biomedical resource; proteins; spectrometry

Using electrospray ionization mass spectrometry (ESI-MS), we have demonstrated the direct observation of NO attached to the heme moiety in horse heart myoglobin (Mb) and in the a- and P- chains of human hemoglobin (Hb). It was found that a narrow range of EST-MS conditions conspire to make observation of Fe-NO interactions challenging, and this is presumably the reason why earlier attempts by other research groups to detect intact Fe-NO products by mass spectrometry were unsuccessful. For Mb and Hb, mass shifts are observed that are consistent with NO modification of the hemoproteins. ESI mass spectra of the apoprotein portions of Mb and Hb in the presence of NO demonstrated the absence of NO modification of the polypeptide backbones. To test the hypothesis that intact nitrosylated heme groups are observable by ESI-MS, a nitrosylated model metalloporphyrin was studied. To test further our hypothesis that the bemoprotein-NO peaks are due to heme nitrosylation and contain no significant contributions from NO modification of the polypeptide backbone, we determined the ESI-MS conditions necessary for observing S-nitrosation of Cys residues in Hb. Our findings demonstrate that (once correct conditions are established) ESI-MS is a powerful tool for the detection of intact Fe-NO interactions in proteins and porphyrins. A paper describing this work is in press in J. Am. Chem. Soc.

使用电喷雾电离质谱仪(ESI-MS) 直接观察到NO附着在血红素部分上 马心脏肌红蛋白(Mb)和人的A-链和P-链 血红蛋白(Hb)。结果表明，EST-MS检测范围较窄。 观察Fe-NO相互作用的条件 具有挑战性，这大概就是为什么早期的尝试 其他研究小组将按质量检测完整的Fe-NO产品 光谱分析都没有成功。对于Mb和Hb，质量移位是 观察到，这与没有修饰的血球蛋白相一致。 日本血吸虫Mb和Hb脱辅基蛋白部分的ESI质谱学研究 NO的存在表明没有修饰 多肽骨架。来检验假设完好无损 用电喷雾质谱(ESI-MS)可以观察到亚硝化的血红素基团 对模型金属卟啉进行了研究。为了进一步检验我们的假设 血蛋白-NO的峰值是由于血红素亚硝化和 不包含重大贡献，因为不修改 多肽骨架，我们确定了电喷雾质谱联用的必要条件 用于观察血红蛋白中半胱氨酸残基的S亚硝化反应。我们的发现 证明(一旦建立正确的条件)ESI-MS是一种 检测完整的Fe-NO相互作用的有力工具 蛋白质和卟啉。描述这项工作的一篇论文正在出版中 J.Am化学。SoC。