Department of Biology, Rensselaer Polytechnic Institute, 110 8th Street, Troy, NY 12180, USA.
J Mol Biol. 2010 Mar 5;396(4):1128-44. doi: 10.1016/j.jmb.2009.12.052. Epub 2010 Jan 4.
Ubiquitin-interacting motifs (UIMs) are an important class of protein domains that interact with ubiquitin or ubiquitin-like proteins. These approximately 20-residue-long domains are found in a variety of ubiquitin receptor proteins and serve as recognition modules towards intracellular targets, which may be individual ubiquitin subunits or polyubiquitin chains attached to a variety of proteins. Previous structural studies of interactions between UIMs and ubiquitin have shown that UIMs adopt an extended structure of a single alpha-helix, containing a hydrophobic surface with a conserved sequence pattern that interacts with key hydrophobic residues on ubiquitin. In light of this large body of structural studies, details regarding the presence and the roles of structural dynamics and plasticity are surprisingly lacking. In order to better understand the structural basis of ubiquitin-UIM recognition, we have characterized changes in the structure and dynamics of ubiquitin upon binding of a UIM domain from the yeast Vps27 protein. The solution structure of a ubiquitin-UIM fusion protein designed to study these interactions is reported here and found to consist of a well-defined ubiquitin core and a bipartite UIM helix. Moreover, we have studied the plasticity of the docking interface, as well as global changes in ubiquitin due to UIM binding at the picoseconds-to-nanoseconds and microseconds-to-milliseconds protein motions by nuclear magnetic resonance relaxation. Changes in generalized-order parameters of amide groups show a distinct trend towards increased structural rigidity at the UIM-ubiquitin interface relative to values determined in unbound ubiquitin. Analysis of (15)N Carr-Purcell-Meiboom-Gill relaxation dispersion measurements suggests the presence of two types of motions: one directly related to the UIM-binding interface and the other induced to distal parts of the protein. This study demonstrates a case where localized interactions among protein domains have global effects on protein motions at timescales ranging from picoseconds to milliseconds.
泛素相互作用基序（UIMs）是一类重要的蛋白质结构域，可与泛素或泛素样蛋白相互作用。这些约 20 个残基长的结构域存在于各种泛素受体蛋白中，作为识别模块，靶向细胞内的靶标，靶标可能是单个泛素亚单位或连接到各种蛋白质的多泛素链。先前对 UIMs 与泛素相互作用的结构研究表明，UIMs 采用单个α-螺旋的伸展结构，包含一个疏水性表面，具有保守的序列模式，与泛素上的关键疏水性残基相互作用。鉴于这些大量的结构研究，关于结构动态和可塑性的存在和作用的细节却出人意料地缺乏。为了更好地理解泛素-UIM 识别的结构基础，我们研究了酵母 Vps27 蛋白的 UIM 结构域结合后泛素结构和动力学的变化。本文报道了一种设计用于研究这些相互作用的泛素-UIM 融合蛋白的溶液结构，发现它由一个定义明确的泛素核心和一个二部分 UIM 螺旋组成。此外，我们通过核磁共振弛豫研究了对接界面的可塑性以及由于 UIM 结合而导致的泛素的全局变化，研究时间范围从皮秒到纳秒和微秒到毫秒的蛋白质运动。酰胺基团的广义顺序参数的变化显示出与未结合的泛素相比，在 UIM-泛素界面处的结构刚性明显增加的趋势。（15）N Carr-Purcell-Meiboom-Gill 弛豫色散测量的分析表明存在两种类型的运动：一种与 UIM 结合界面直接相关，另一种诱导至蛋白质的远端部分。这项研究证明了在蛋白质结构域之间的局部相互作用在皮秒到毫秒的时间范围内对蛋白质运动具有全局影响的情况。
