Biocatalytic asymmetric rearrangement of a methylene-interrupted bis-epoxide: simultaneous control of four asymmetric centers through a biomimetic reaction cascade

Asymmetric enzyme-catalyzed hydrolysis of methylene-interrupted bis-epoxides 1 a and 1 b catalyzed by bacterial epoxide hydrolases furnished tetrahydrofuran derivatives 2 a and 2 b through a hydrolysis-rearrangement cascade. Whereas racemic bis-oxiranes 1 b-d underwent kinetic resolution with moderate stereoselectivities to yield products with up to 92 % ee and 66 % de: meso-bis-oxirane cis,cis-1 a was transformed into (6R,7R,9S,10S)-2 a in 94 % ee and 89 % de at high conversion (85 %) by Rhodococcus sp. CBS 717.73 as the major product. The reaction sequence resembles a biomimetic reaction cascade and provides an efficient entry into the structural core of annonaceous acetogenins with simultaneous control of four stereocenters.     

反相高效液相色谱分离黄酮类化合物的研究

根据18种黄酮类化合物在C_3、C_(18)和CN三种固定相上的梯度洗脱反相高效液相色谱分离的研究,建立了一种连续快速分离植物提取物中黄酮类化合物的方法。此法可在55 min内完成一个样品的分离及柱平衡。最佳波长和温度分别是280nm和35℃。     

Generation and Use of Glycosyl Radicals under Acidic Conditions: Glycosyl Sulfinates as Precursors

We herein report a method that enables the generation of glycosyl radicals under highly acidic conditions. Key to the success is the design and use of glycosyl sulfinates as radical precursors, which are bench-stable solids and can be readily prepared from commercial starting materials. This development allows the installation of glycosyl units onto pyridine rings directly by the Minisci reaction. We further demonstrate the utility of this method in the late-stage modification of complex drug molecules, including the anticancer agent camptothecin. Experimental studies provide insight into the reaction mechanism.     

绿豆环氧水解酶催化对硝基苯乙烯氧化物的对映归一性水解

利用绿豆环氧水解酶催化(R,S)-对硝基苯乙烯氧化物水解,产物(R)-对硝基苯乙二醇的产率超过常规酶促拆分的极限产率(50%), 表明发生了对映体会聚. 采用绿豆粉剂和粗酶制剂考察了不同条件下的生物催化反应,结果表明,添加吐温-80 作为分散剂对反应有促进作用. 对该酶的固定化进行了初步研究,发现酶经硅藻土吸附固定化后稳定性显著增加. 在克级规模制备实验中,产物的总产率达73.3%;产物经一步重结晶,得到了光学纯度大于99%的(R)-对硝基苯乙二醇.     

High-performance liquid chromatography quantitation of limonin D-ring lactone hydrolase and limonoate dehydrogenase activities

Ion-pairing reversed-phase HPLC has been used to separate and determine different derivatives of limonin, i.e. disodium limonoate, 17-dehydrolimonoate, limonoate A-ring lactone and 17-dehydrolimonoate A-ring lactone. Consequently, it has been possible to measure, by this chromatographic technique, the activities of the two enzymes that sequentially catalize the first two steps in limonin degradation: limonin D-ring lactone hydrolase and limonoate dehydrogenase. In both cases linear responses were found when using increasing enzyme amounts. The determination of limonoate dehydrogenase by HPLC measurements of limonoid products was compared with that obtained by the spectrophotometric method, based on pyridine nucleotides reduction. A good correlation was detected by using both procedures, which allows the proposition of the chromatographic method as an alternative to pyridine nucleotides determination. Besides, the chromatographic method appears to be a better assay than thin-layer chromatography or radioactive procedures, the methods traditionally used to measure limonin D-ring lactone hydrolase activity.     

Natural Product Inhibition and Enzyme Kinetics Related to Phylogenetic Characterization for Bacterial Peptidyl-tRNA Hydrolase 1

With the relentless development of drug resistance and re-emergence of many pathogenic bacteria, the need for new antibiotics and new antibiotic targets is urgent and growing. Bacterial peptidyl-tRNA hydrolase, Pth1, is emerging as a promising new target for antibiotic development. From the conserved core and high degree of structural similarity, broad-spectrum inhibition is postulated. However, Pth1 small-molecule inhibition is still in the earliest stages. Focusing on pathogenic bacteria, herein we report the phylogenetic classification of Pth1 and natural product inhibition spanning phylogenetic space. While broad-spectrum inhibition is found, narrow-spectrum and even potentially clade-specific inhibition is more frequently observed. Additionally reported are enzyme kinetics and general in vitro Pth1 solubility that follow phylogenetic boundaries along with identification of key residues in the gate loop region that appear to govern both. The studies presented here demonstrate the sizeable potential for small-molecule inhibition of Pth1, improve understanding of Pth enzymes, and advance Pth1 as a much-needed novel antibiotic target.     

Glycosyl‐Substituted Dicarboxylates as Detergents for the Extraction,Overstabilization, and Crystallization of Membrane Proteins

To tackle the problems associated with membrane protein (MP) instability in detergent solutions, we designed a series of glycosyl‐substituted dicarboxylate detergents (DCODs) in which we optimized the polar head to clamp the membrane domain by including, on one side, two carboxyl groups that form salt bridges with basic residues abundant at the membrane–cytoplasm interface of MPs and, on the other side, a sugar to form hydrogen bonds. Upon extraction, the DCODs 8 b , 8 c , and 9 b preserved the ATPase function of BmrA, an ATP‐binding cassette pump, much more efficiently than reference or recently designed detergents. The DCODs 8 a , 8 b , 8 f , 9 a , and 9 b induced thermal shifts of 20 to 29 °C for BmrA and of 13 to 21 °C for the native version of the G‐protein‐coupled adenosine receptor A_2AR. Compounds 8 f and 8 g improved the diffraction resolution of BmrA crystals from 6 to 4 Å. DCODs are therefore considered to be promising and powerful tools for the structural biology of MPs.