Optimized Biocatalytic Synthesis of 2-Selenopyrimidine Nucleosides by Transglycosylation**

Selenium-modified nucleosides are powerful tools to study the structure and function of nucleic acids and their protein interactions. The widespread application of 2-selenopyrimidine nucleosides is currently limited by low yields in established synthetic routes. Herein, we describe the optimization of the synthesis of 2-Se-uridine and 2-Se-thymidine derivatives by thermostable nucleoside phosphorylases in transglycosylation reactions using natural uridine or thymidine as sugar donors. Reactions were performed at 60 or 80 °C and at pH 9 under hypoxic conditions to improve the solubility and stability of the 2-Se-nucleobases in aqueous media. To optimize the conversion, the reaction equilibria in analytical transglycosylation reactions were studied. The equilibrium constants of phosphorolysis of the 2-Se-pyrimidines were between 5 and 10, and therefore differ by an order of magnitude from the equilibrium constants of any other known case. Hence, the thermodynamic properties of the target nucleosides are inherently unfavorable, and this complicates their synthesis significantly. A tenfold excess of sugar donor was needed to achieve 40−48 % conversion to the target nucleoside. Scale-up of the optimized conditions provided four Se-containing nucleosides in 6–40 % isolated yield, which compares favorably to established chemical routes.     

8-R-9苄基-9H-嘌呤衍生物的合成研究

采用 (Ph3 P) 2 PdCl2 为催化剂 ,DMF为溶剂 ,对 8 碘 9 苄基 9H 嘌呤与有机锡试剂RSnBu3 (R=乙烯基、2 噻吩基、2 呋喃基、苯乙炔基和苯基等 )之间的Stille偶合反应进行了研究 ,合成出了5种 8位取代的嘌呤衍生物。在反应温度为 80℃ ,n(8 碘 9 苄基 9H 嘌呤 )∶n (RSnBu3 )∶n〔(Ph3 P) 2 PdCl2 〕 =1 0∶1 2∶0 0 5的较佳工艺条件下 ,产品收率 4 1%～ 91%。用1H NMR、13 C NMR和MS对产物进行了表征     

Uric Acid: The Sperm-Release Pheromone of the Marine Polychaete Platynereis dumerilii

In the marine polychaete, Platynereis dumerilii, reproductive behavior in the two sexes is synchronized by the consecutive discharge of male and female sex-specific pheromones. After the female releases the eggs into the free water column, immediate fertilization is achieved by several males circling around the eggs emitting sperm clouds. We report the isolation and identification of the sperm-release pheromone present in the coelomic fluid of sexually mature females. Each step in isolation was guided by bioassay. Isolation methods included extraction and solvent partitioning and separation methods included ultrafiltration and high-performance liquid chromatography. Uric acid was identified as the sperm-release pheromone that is discharged by the female with release of the eggs. The threshold concentration for sperm release by males was determined as 0.6 M.     

一类核苷类似物关键中间体的合成

对一类核苷类似物的关键中间体———(4R,5R) 4 (N 甲基羟氨基) 5 [(叔丁基二苯基硅烷基)氧甲基] 3,4 二氢 2(5H) 呋喃酮的合成进行了研究。以L 抗坏血酸为原料,经Pd/C催化加氢和缩酮保护,生成5,6 O 异亚丙基 L 古洛糖酸 1,4 内酯,产率74 3%。该内酯经NaIO4氧化、Wittig反应、水解成环和柱色谱分离,得(R) (+) 5 羟甲基 2 (5H) 呋喃酮,产率43 0%。最后该呋喃酮再经硅烷保护和羟胺Michael加成,即得到目标化合物。这7步反应的总产率27 9%。     

8-碘-9-四氢吡喃-2-基-9H-嘌呤的合成

以 9-四氢吡喃 - 2 -基 - 9H-嘌呤、丁基锂和 I2 等为主要原料 ,四氢呋喃 ( THF)为溶剂 ,合成了 8-碘 - 9-四氢吡喃 - 2 -基 - 9H-嘌呤。考察了主要反应条件对反应的影响 ,确定了较佳工艺条件 :反应温度 - 78℃ ,n( 9-四氢吡喃 - 2 -基 - 9H-嘌呤 )∶ n(丁基锂 )∶ n(碘 ) =1 .0∶ 1 .5∶ 1 .5 ,反应时间 2 h。此条件下 ,产物收率 >90 %。同时 ,采用 1H- NMR、13C-NMR和 MS对产物结构进行了表征     

Milk feeding level and starter protein content: Effects on growth performance,blood metabolites,and urinary purine derivatives of Holstein dairy calves

The objective of this study was to investigate the effects of milk allowances equal to 526 g/d as moderate (MOD) versus 790 g/d of milk dry matter as high (HI), and starter diets containing 18% or 23% crude protein (CP), on growth performance, blood metabolites, and purine derivative (PD) excretion in the urine of dairy calves. A total of 52 female Holstein dairy calves (40.8 kg of body weight) were randomly assigned to the experimental diets. The treatments were (1) moderate milk and 18% CP starter diet (MOD-18CP); (2) MOD and 23% CP starter diet (MOD-23CP); (3) high milk and 18% CP starter diet (HI-18CP); and (4) HI and 23% CP starter diet (HI-23CP). Calves had free access to a starter feed and water and were weaned on d 53 but remained in the study until d 73. Urine samples were collected during the preweaning period (for 6 consecutive days between d 35 and 40) and postweaning period (for 6 consecutive days between d 65 and 70) to investigate urinary excretion of PD. Starter feed intake, β-hydroxybutyrate (BHB), and blood urea concentrations were reduced; however, average daily gain (ADG) and blood glucose levels increased in calves fed HI before weaning compared with MOD. During the preweaning period, high milk feeding increased total urinary PD excretion but decreased it after weaning. The 23CP diet resulted in higher feed intake and ADG before weaning and higher excretion of allantoin and total excretion of PD compared with the 18CP diet. The HI-23CP treatment resulted in the greatest withers and hip heights at weaning and final measurement, as well as the highest preweaning blood insulin concentrations. In terms of rumen development, MOD-23CP showed the greatest benefits based on starter intake, blood BHB concentration, and urinary excretion of PD. Based on the higher urinary excretion of PD found in HI-fed calves before weaning, it is possible that milk feeding overestimates estimated microbial yield. The results suggest that feeding starters with a higher proportion of CP may help maintain a more balanced ratio of CP to ME during high milk feeding, to avoid protein deficiency due to low starter intake. When calves are fed a high milk allowance, urine excretion of PD may be misinterpreted as a measure of estimated microbial growth and rumen development; this should be considered during calculations of estimated microbial yield in milk-fed calves.     

Nucleoside‐Based Diarylethene Photoswitches: Synthesis and Photochromic Properties

Diarylethene photoswitches based on the natural nucleoside deoxyadenosine were designed and synthesized. In aqueous solution, some of them exhibited good photochromic properties, including clear changes in color upon irradiation at 365 nm, red‐shifts of the absorption wavelength, with good fatigue resistance, thermal stability, conversion efficiency, and base‐pairing properties.     

卡培他滨为基础一线方案治疗Her2阴性转移性乳腺癌的单中心回顾性研究及TYMP基因多态性分析

目的:本研究旨在探讨卡培他滨为基础一线方案治疗Her2阴性转移性乳腺癌的单中心回顾性研究及TYMP基因多态性分析。方法:本研究为回顾性分析,入组患者为2010年1月到2017年12月期间在首都医科大学附属北京天坛医院乳腺科和肿瘤科确诊为Her2阴性的且接受卡培他滨为基础一线治疗的转移性乳腺癌患者共215例。收集患者的外周血液进行基因分型,并进行mRNA表达测定和相关性分析,电话随访获取患者预后数据。用Kaplan-Meier生存分析方法分析不同基因型和预后的关联,用Cox模型对其他变量进行校正。结果:纳入分析的TYMP的遗传变异位点经过NCBI数据库查阅在中国人群中突变频率大于10%,在预后分析上发现1412C>T位点的关联性。1412C>T位点在本研究中的突变频率为：CC基因型137例（63.72%）,CT基因型70例（32.56%）,TT基因型8例（3.72%）,最小等位基因频率为0.20,符合HW平衡（P=0.798）。以显性遗传的方式把CT和TT基因型合并,对两组基因型患者进行预后分析发现：CT/TT和CC基因型患者的客观缓解率（ORR）分别为56.41%和40.15%（P=0.021）,疾病控制率分别为85.90%和72.99%（P=0.029）,中位无进展生存期（mPFS）分别为11.5和7.6个月（P=0.001）。另外,两组基因型患者的中位总生存期（mOS）分别为26.6和24.7个月（P=0.022）。针对PFS构建多变量的Cox模型校正后,CT/TT基因型仍然是PFS的独立影响因素（OR=0.56,P=0.011）。87例癌组织标本的mRNA表达分析结果表明CT/TT基因型患者相对于CC型患者,癌组织中TYMP的mRNA表达明显较高（P<0.001）。结论:TYMP基因1412C>T位点是卡培他滨作为基础化疗药物在一线治疗Her2阴性转移性乳腺癌患者疗效的预测因子。     

A Toxoplasma gondii Oxopurine Transporter Binds Nucleobases and Nucleosides Using Different Binding Modes

Toxoplasma gondii is unable to synthesize purines de novo, instead salvages them from its environment, inside the host cell, for which they need high affinity carriers. Here, we report the expression of a T. gondii Equilibrative Nucleoside Transporter, Tg244440, in a Trypanosoma brucei strain from which nucleobase transporters have been deleted. Tg244440 transported hypoxanthine and guanine with similar affinity (K_m ~1 µM), while inosine and guanosine displayed K_i values of 4.05 and 3.30 µM, respectively. Low affinity was observed for adenosine, adenine, and pyrimidines, classifying Tg244440 as a high affinity oxopurine transporter. Purine analogues were used to probe the substrate-transporter binding interactions, culminating in quantitative models showing different binding modes for oxopurine bases, oxopurine nucleosides, and adenosine. Hypoxanthine and guanine interacted through protonated N1 and N9, and through unprotonated N3 and N7 of the purine ring, whereas inosine and guanosine mostly employed the ribose hydroxy groups for binding, in addition to N1H of the nucleobase. Conversely, the ribose moiety of adenosine barely made any contribution to binding. Tg244440 is the first gene identified to encode a high affinity oxopurine transporter in T. gondii and, to the best of our knowledge, the first purine transporter to employ different binding modes for nucleosides and nucleobases.     

Effect of Betaine and Arginine on Interaction of αB-Crystallin with Glycogen Phosphorylase b

Protein–protein interactions (PPIs) play an important role in many biological processes in a living cell. Among them chaperone–client interactions are the most important. In this work PPIs of αB-crystallin and glycogen phosphorylase b (Phb) in the presence of betaine (Bet) and arginine (Arg) at 48 °C and ionic strength of 0.15 M were studied using methods of dynamic light scattering, differential scanning calorimetry, and analytical ultracentrifugation. It was shown that Bet enhanced, while Arg reduced both the stability of αB-crystallin and its adsorption capacity (AC₀) to the target protein at the stage of aggregate growth. Thus, the anti-aggregation activity of αB-crystallin increased in the presence of Bet and decreased under the influence of Arg, which resulted in inhibition or acceleration of Phb aggregation, respectively. Our data show that chemical chaperones can influence the tertiary and quaternary structure of both the target protein and the protein chaperone. The presence of the substrate protein also affects the quaternary structure of αB-crystallin, causing its disassembly. This is inextricably linked to the anti-aggregation activity of αB-crystallin, which in turn affects its PPI with the target protein. Thus, our studies contribute to understanding the mechanism of interaction between chaperones and proteins.