Masterson Larry R, Tonelli Marco, Markley John L, Veglia Gianluigi
Department of Chemistry and Biochemistry, University of Minnesota, Minneapolis, Minnesota 55455-0431, USA.
J Am Chem Soc. 2008 Jun 25;130(25):7818-9. doi: 10.1021/ja802701w. Epub 2008 May 31.
We present a procedure for isolating subspectra corresponding to individual protein or peptide components in a ternary mixture or complex. Each of the three-component species is labeled differently: species A uniformly with 15N, species B uniformly with 15N and 13C, and species C uniformly with 15N but selectively with 13C' or 13Calpha. By using the dual carbon label selective HSQC (DCLS-HSQC) pulse sequence and exploiting differences in 1J 15N-13C coupling patterns to filter selected 15N resonances from detection during a constant time period, a subspectrum from each species can be generated from three spectra acquired from a single sample. Many important biological pathways involve dynamic interactions among members of multicomponent protein assemblies, and this approach offers a powerful way to monitor such processes.
我们提出了一种从三元混合物或复合物中分离出对应于单个蛋白质或肽成分的子光谱的方法。三种成分的每种都用不同的方式标记：成分A用15N均匀标记，成分B用15N和13C均匀标记，成分C用15N均匀标记但用13C'或13Calpha选择性标记。通过使用双碳标记选择性HSQC（DCLS-HSQC）脉冲序列，并利用1J 15N-13C耦合模式的差异，在恒定时间段内从检测中过滤选定的15N共振，可以从单个样品获取的三个光谱中生成每个成分的子光谱。许多重要的生物途径涉及多成分蛋白质组装体成员之间的动态相互作用，这种方法为监测此类过程提供了一种强大的方式。
