Stinson Craig A, Xia Yu
Department of Chemistry, Purdue University, West Lafayette, IN, 47907-2084, USA.
J Am Soc Mass Spectrom. 2014 Jul;25(7):1192-201. doi: 10.1007/s13361-014-0898-8. Epub 2014 May 3.
In biological systems, carbon-centered small molecule radicals are primarily formed via external radiation or internal radical reactions. These radical species can react with a variety of biomolecules, most notably nucleic acids, the consequence of which has possible links to gene mutation and cancer. Sulfur-containing peptides and proteins are reactive toward a variety of radical species and many of them behave as radical scavengers. In this study, the reactions between alkyl alcohol carbon-centered radicals (e.g., •CH2OH for methanol) and cysteinyl peptides within a nanoelectrospray ionization (nanoESI) plume were explored. The reaction system involved ultraviolet (UV) irradiation of a nanoESI plume using a low pressure mercury lamp consisting of 185 and 254 nm emission bands. The alkyl alcohol was added as solvent into the nanoESI solution and served as the precursor of hydroxyalkyl radicals upon UV irradiation. The hydroxyalkyl radicals subsequently reacted with cysteinyl peptides either containing a disulfide linkage or free thiol, which led to the formation of peptide-S-hydroxyalkyl product. This radical reaction coupled with subsequent MS/MS was shown to have analytical potential by cleaving intrachain disulfide linked peptides prior to CID to enhance sequence information. Tandem mass spectrometry via collision-induced dissociation (CID), stable isotope labeling, and accurate mass measurement were employed to verify the identities of the reaction products.
在生物系统中，以碳为中心的小分子自由基主要通过外部辐射或内部自由基反应形成。这些自由基物种可与多种生物分子发生反应，最显著的是核酸，其结果可能与基因突变和癌症有关。含硫肽和蛋白质对多种自由基物种具有反应性，其中许多表现为自由基清除剂。在本研究中，探索了纳米电喷雾电离（nanoESI）羽流中烷基醇碳中心自由基（例如，甲醇的•CH2OH）与半胱氨酰肽之间的反应。反应体系涉及使用由185和254 nm发射带组成的低压汞灯对nanoESI羽流进行紫外线（UV）照射。将烷基醇作为溶剂添加到nanoESI溶液中，并在UV照射下作为羟烷基自由基的前体。羟烷基自由基随后与含有二硫键或游离巯基的半胱氨酰肽发生反应，从而导致形成肽-S-羟烷基产物。通过在碰撞诱导解离（CID）之前切割链内二硫键连接的肽以增强序列信息，这种自由基反应与随后的MS/MS相结合显示出具有分析潜力。采用通过碰撞诱导解离（CID）的串联质谱、稳定同位素标记和精确质量测量来验证反应产物的身份。
