文章基本信息

标题：The structure of the catechin-binding site of human sulfotransferase 1A1
本地全文：下载
作者：Ian Cook ; Ting Wang ; Mark Girvin 等
期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2016
卷号：113
期号：50
页码：14312-14317
DOI：10.1073/pnas.1613913113
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceThe human cytosolic sulfotransferases (SULTs) regulate scores of disease-relevant cellular processes through site-specific attachment of the sulfuryl-group (-SO3) to signaling small molecules. Recent work reveals that SULT1A1, the predominant SULT in adult liver, harbors two allosteric-binding pockets, each of which binds a separate and complex class of biomorphic xenobiotics. These sites represent "portals" through which SULTs communicate with their environments, and their structures reveal the molecular basis of this dialog and provide templates for identifying new allosteres. Here, we present a structure of a SULT allosteric site--the catechin-binding site of SULT1A--which was determined using a spin-label triangulation method. We are just beginning to understand the allosteric regulation of the human cytosolic sulfotransferase (SULTs) family--13 disease-relevant enzymes that regulate the activities of hundreds, if not thousands, of signaling small molecules. SULT1A1, the predominant isoform in adult liver, harbors two noninteracting allosteric sites, each of which binds a different molecular family: the catechins (naturally occurring flavonols) and nonsteroidal antiinflammatory drugs (NSAIDs). Here, we present the structure of an SULT allosteric binding site--the catechin-binding site of SULT1A1 bound to epigallocatechin gallate (EGCG). The allosteric pocket resides in a dynamic region of the protein that enables EGCG to control opening and closure of the enzymes active-site cap. Furthermore, the structure offers a molecular explanation for the isozyme specificity of EGCG, which is corroborated experimentally. The binding-site structure was obtained without X-ray crystallography or multidimensional NMR. Instead, a SULT1A1 apoprotein structure was used to guide positioning of a small number of spin-labeled single-Cys mutants that coat the entire enzyme surface with a paramagnetic field of sufficient strength to determine its contribution to the bound ligands transverse (T2) relaxation from its 1D solution spectrum. EGCG protons were mapped to the protein surface by triangulation using the T2 values to calculate their distances to a trio of spin-labeled Cys mutants. The final structure was obtained using distance-constrained molecular dynamics docking. This approach, which is readily extensible to other systems, is applicable over a wide range of ligand affinities, requires little protein, avoids the need for isotopically labeled protein, and has no protein molecular weight limitations.
关键词：sulfotransferase ; structure ; NMR ; allostery ; catechin