使用非特异性蛋白酶K消化和CID可裂解交联分析方法对Orbitrap LC/ESI-MS/MS数据进行二硫键的全面鉴定。
Comprehensive identification of disulfide bonds using non-specific proteinase K digestion and CID-cleavable crosslinking analysis methodology for Orbitrap LC/ESI-MS/MS data.
作者信息
Makepeace Karl A T, Serpa Jason J, Petrotchenko Evgeniy V, Borchers Christoph H
机构信息
University of Victoria - Genome British Columbia Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada.
University of Victoria - Genome British Columbia Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada; Department of Biochemistry & Microbiology, University of Victoria, Victoria, British Columbia V8P 5C2, Canada.
出版信息
Methods. 2015 Nov 1;89:74-8. doi: 10.1016/j.ymeth.2015.02.021. Epub 2015 Mar 6.
Disulfide bonds are valuable constraints in protein structure modeling. The Cys-Cys disulfide bond undergoes specific fragmentation under CID and, therefore, can be considered as a CID-cleavable crosslink. We have recently reported on the benefits of using non-specific digestion with proteinase K for inter-peptide crosslink determination. Here, we describe an updated application of our CID-cleavable crosslink analysis software and our crosslinking analysis with non-specific digestion methodology for the robust and comprehensive determination of disulfide bonds in proteins, using Orbitrap LC/ESI-MS/MS data.
二硫键是蛋白质结构建模中有价值的限制因素。半胱氨酸-半胱氨酸二硫键在碰撞诱导解离（CID）下会发生特异性断裂，因此可被视为一种可被CID裂解的交联键。我们最近报道了使用蛋白酶K进行非特异性消化以确定肽间交联的益处。在此，我们描述了我们的CID可裂解交联分析软件的更新应用，以及使用非特异性消化方法进行交联分析，以利用轨道阱液相色谱/电喷雾串联质谱（Orbitrap LC/ESI-MS/MS）数据对蛋白质中的二硫键进行可靠而全面的测定。
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