Structural analysis of a bacterial UDP-sugar 2-epimerase reveals the active site architecture before and after catalysis

James B. Thoden, James O. Mcknight, Charles W. Kroft, Joshua D. T. Jast,Hazel M. Holden

JOURNAL OF BIOLOGICAL CHEMISTRY(2023)

引用 0|浏览3
暂无评分
摘要
The sugar, 2,3-diacetamido-2,3-dideoxy-D-mannuronic acid, was first identified X40 years ago in the O-antigen of Pseudomonas aeruginosa O:3,a,d. Since then, it has been observed on the O-antigens of various pathogenic Gram -negative bacteria including Bordetella pertussis, Escherichia albertii, and Pseudomonas mediterranea. Previous studies have established that five enzymes are required for its biosynthesis beginning with uridine dinucleotide (UDP)-N-acetyl-D-glucosamine (UDPGlcNAc). The final step in the pathway is catalyzed by a 2epimerase, which utilizes UDP-2,3-diacetamido-2,3-dideoxy-D- glucuronic acid as its substrate. Curious as to whether this biochemical pathway is found in extreme thermophiles, we examined the published genome sequence for Thermus thermophilus HB27 and identified five ORFs that could possibly encode for the required enzymes. The focus of this investigation is on the ORF WP_011172736, which we demonstrate encodes for a 2-epimerase. For this investigation, ten high resolution Xray crystallographic structures were determined to resolutions of 2.3 angstrom or higher. The models have revealed the manner in which the 2-epimerase anchors its UDP-sugar substrate as well as its UDP-sugar product into the active site. In addition, this study reveals for the first time the manner in which any sugar 2epimerase can simultaneously bind UDP-sugars in both the active site and the allosteric binding region. We have also demonstrated that the T. thermophilus enzyme is allosterically regulated by UDP-GlcNAc. Whereas the sugar 2-epimerases that function on UDP-GlcNAc have been the focus of past biochemical and structural analyses, this is the first detailed investigation of a 2-epimerase that specifically utilizes UDP-2,3diacetamido-2,3-dideoxy-D-glucuronic acid as its substrate.
更多
查看译文
关键词
catalysis,udp-sugar
AI 理解论文
溯源树
样例
生成溯源树,研究论文发展脉络
Chat Paper
正在生成论文摘要