hKv4.3基因5′非翻译区序列S160功能分析

hKv4.3基因是形成瞬时外向钾电流Ito的主要分子基础,它在心脏和神经细胞中大量表达,但在其它组织中则未见大量表达.为了研究hKv4.3表达在基因水平的调节,将hKv4.3基因的5′非翻译区的一段序列( 2~ 160,称之为S160)克隆到报告质粒中,进行瞬时表达.发现S160对hKv4.3基因的启动子和SV40的启动子都有强烈的抑制作用,没有方向特异性,但却有位置特异性.经删除突变分析,在S160片段中发现了一个抑制元件S(GAGGGGTTAA),它位于hKv4.3基因中转录起始位点下游20~30bp处.在此基础上,用RT-PCR方法对mRNA进行定量分析,初步确认这个抑制元件对蛋白表达的抑制过程是在翻译水平上.     

hKv4.3基因5'非翻译区序列S160功能分析

hKv4.3基因是形成瞬时外向钾电流Ito的主要分子基础, 它在心脏和神经细胞中大量表达, 但在其它组织中则未见大量表达. 为了研究hKv4.3表达在基因水平的调节, 将hKv4.3基因的5'非翻译区的一段序列(+2~+160, 称之为S160)克隆到报告质粒中, 进行瞬时表达. 发现S160对hKv4.3基因的启动子和SV40的启动子都有强烈的抑制作用, 没有方向特异性, 但却有位置特异性. 经删除突变分析, 在S160片段中发现了一个抑制元件S(GAGGGGTTAA), 它位于hKv4.3基因中转录起始位点下游20-30 bp处. 在此基础上, 用RT-PCR方法对mRNA进行定量分析, 初步确认这个抑制元件对蛋白表达的抑制过程是在翻译水平上.     

hKv4.3基因5＇非翻译区序列S160功能分析

hKv4.3基因是形成瞬时外向钾电流Ito的主要分子基础,它在心脏和神经细胞中大量表达,但在其它组织中则未见大量表达。为了研究hKv4.3表达在基因水平的调节,将hKv4.3基因的5＇非翻译区的一段序列（＋2～＋160,称之为S160）克隆到报告质粒中,进行瞬时表达。发现S160对hKv4.3基因的启动子和SV40的启动子都有强烈的抑制作用,没有方向特异性,但却有位置特异性。经删除突变分析,在S160片段中发现了一个抑制元件S（GAGGGGTTAA）,它位于hKv4.3基因中转录起始位点下游20～30 bp处。在此基础上,用RT-PCR方法对mRNA进行定量分析,初步确认这个抑制元件对蛋白表达的抑制过程是在翻译水平上。     

Synthesis of 5′-deoxy-5′-[18F]fluoro-adenosine by Radiofluorination of 5′-deoxy-5′-haloadenosine Derivatives

5-Deoxy-5-[¹⁸F]fluoro-adenosine was synthesised by nucleophilic radiofluorination reactions of 5-deoxy-5-haloadenosines. The homogeneous isotope exchange in 5-deoxy-5-fluoro-adenosine was also investigated. The conversion of these reactions was found to be rather low and depends on the strength of the halogen-carbon bond: 0.248% for chloride-, 0.488% for bromide- and 1.070% for iodide-derivative; there was no reaction observed in the case of fluoro-compound.     

Syntheses of 5′-O-glycosylnucleosides

A general synthetic approach to 2,3-unsaturated glycosides connecting with nucleosides involving Ferrier rearrangements of glycals is discussed. The new compounds were identified by NMR and MS (HRFAB⁺). The hydroxylation of the resulting 2,3-unsaturated glycosides was completed using OsO₄ to give 5′-O-glycosylnucleosides in good yield.     

Chemical 5′-phosphorylation of oligonucleotidesvaluable in automated DNA synthesis

A new method has been developed for the chemical 5′-phosphorylation ofoligonucleotides starting from easily available phosphorous-III compounds using phosphate protecting groups which can be cleaved off via β-elimination. The hydrophobic 2-(p-nitrophenyl)-ethyl group, at the phosphodiester level can serve as a purification handle in reversed phase HPLC. Moreover, the procedure has been adapted to automated DNA synthesis.     

Study of Phase Transformation of Guanosine 5′-Monophosphate in Drowning-Out Crystallization

The present study used a mechanistic approach to control the phase transformation of guanosine 5′-monophosphate (GMP) via the operating conditions of agitation and feed concentration during drowning-out crystallization. First, Fourier transform infrared and UV/vis spectrophotometry were successfully applied to monitor the mass fraction of GMP polymorphs (amorphous and hydrate crystalline GMPs) and GMP supersaturation, respectively, during the crystallization. The phase transformation of amorphous GMP into hydrate crystals was significantly influenced by the agitation, which promoted the mass transfer of GMP dissolution and growth. Therefore, the phase transformation was quickly finished when increasing the agitation speed. However, a high agitation caused breakage of the hydrate crystals, resulting in a reduced crystal size with a bimodal distribution. The phase transformation was also influenced by the GMP feed concentration, as the crystal growth was promoted and the crystal size increased when increasing the feed concentration up to 61 g/l. However, a further increase in the feed concentration caused secondary nucleation due to the induction of a high supersaturation level during the phase transformation, leading to a small crystal size with a bimodal distribution. In addition, the rectangular-shaped hydrate GMP crystals exhibited a higher growth rate in the b direction rather than the a direction. Therefore, the crystal morphology shifted from a long rectangle to a square when increasing the feed concentration.     

Thermal analysis of 5′-deoxy-5′-iodo-2′,3′-O-isopropylidene-5-fluorouridine

The melting temperature, melting enthalpy, and specific heat capacities (C _p) of 5′-deoxy-5′-iodo-2′,3′-O-isopropylidene-5-fluorouridine (DIOIPF) were measured using DSC-60 Differential Scanning Calorimetry. The melting temperature and melting enthalpy were obtained to be 453.80 K and 33.22 J g^?1, respectively. The relationship between the specific heat capacity and temperature was obtained to be C _p/J g^?1 K^?1 = 2.0261 – 0.0096T + 2 × 10^?5 T ² at the temperature range from 320.15 to 430.15 K. The thermal decomposition process was studied by the TG–DTA analyzer. The results showed that the thermal decomposition temperature of DIOIPF was above 487.84 K, and the decomposition process can be divided into three stages: the first stage is the decomposition of impurities, the mass loss in the second stage may be the sublimation of iodine and thermal decomposition process of the side-group C₄H₂O₂N₂F, and the third stage may be the thermal decomposition process of both the groups –CH₃ and –CH₂OCH₂–. The obtained thermodynamic basic data are helpful for exploiting new synthetic method, engineering design, and commercial process of DIOIPF.     

Synthesis of 2′-5′,3′-5′ linked triadenylates

2′-5′,3′-5′ Linked triadenylates have been synthesized by direct bisadenylylation of adenosine 2′ and 3′ hydroxyls with an adenosine 5′-phosphorochloridite followed by oxidation.     

Protonation Constants of Uridine 5′-Monophosphate in Different Aqueous Solutions of Methanol

The protonation equilibria of uridine 5′-monophosphate disodium salt (UMP) was determined in binary solvent mixtures of water–methanol containing 0, 10, 15, 20, 25, 30, 35, 40, 45, and 50 % (v/v) methanol, using a combination of spectrophotometric and potentiometric methods at 25 °C and constant ionic strength (0.1 mol·dm^?3 NaClO₄). The protonation constants were analyzed using Kamlet, Abboud, and Taft parameters. A good linear correlation of the protonation constants (on the logarithmic scale) was obtained. Dual-parameter correlation of log₁₀ K versus π* (dipolarity/polarizability) and α (hydrogen-bond donor acidity), as well as π* and β (hydrogen-bond acceptor basicity), gave good results in various aqueous solutions of methanol. Finally, the results are compared with CMP, a homolog of UMP, and are discussed in terms of the effect of the solvent on the protonation constants.     

A multinuclear NMR study on the microscopic ionization constants of adenosine-5′-triphosphate in aqueous solution

A ¹H, ¹³C and ³¹P NMR study of adenosine-5′-triphosphate has been performed as a function of pH. The examination of the chemical shifts allowed the proposal of an ionization scheme in which the third deprotonation takes place both on the adenine ring and on the phosphate group in a ratio of about 2:1. Furthermore, an analysis of carbon relaxation times supports this mechanism of ionization and confirms that the adenine ring binds the proton both on the NH₂ group and the N₇ nitrogen atom.     

Synthesis of Isonucleoside 5′-Triphosphates

Abstract

A series of isonucleoside 5′-triphosphates were synthesized via a rapid and efficient method. The structures of the triphosphates (8a–8d) were characterized by ³¹P NMR and TOF-MS.     

Stereoselective synthesis of 5′-hydroxyzearalenone

A first stereoselective total synthesis of 14-memebered β-resorcylic macrolactone 5′-hydroxyzearalenone (1) has been achieved. The key steps are Jocobsen hydrolytic kinetic resolution, Sharpless asymmetric dihydroxylation, Vilsmeier-Haack reaction, Mitsunobu esterification and ring-closing metathesis.     

干扰素的功能表达机理——Ⅹ.ATP对ppp A2′p5′A2′p5′A受体的拮抗作用

pppA2′p5′A2′p5′A(2′-5′P_3A_3)受体可能是第一个寡核苷酸受体。但ATP能与2′-5′P_3A_3竞争此受体,那么ATP的作用是什么呢?本文证明ATP虽然对巨噬细胞的结合具有饱和性和可逆性,似乎是2′-5′P_3A_3受体的一个配基,但ATP并不能提高巨噬细胞的吞噬功能,而且当ATP和2′-5′P_3A_3同时作用于巨噬细胞时,ATP反而抑制2′-5′P_3A_3~′对巨噬细胞的生物效应。如果ATP先与巨噬细胞作用占据了2′-5′P_3A_3所要结合的位点后,再加2′-5′P_3A_3时,则2′-5′P_3A_3的结合能力和引起的生物学效应都明显地被阻断了,这表明,2′-5′P_3A_3的作用是通过它的受体而发挥的,ATP则可能是2′-5′P_3A_3受体的一个拮抗剂。     

2′-5′连接寡聚核苷酸——2′-5′A核的固相合成

本文用磷酸三酯法,在交联聚丙烯酰吗福啉载体上,以1,3,5-三甲苯磺酰氯(MS)加N-甲基咪唑(N-Mein)为缩合剂,合成了2′-5′相连的寡聚腺苷酸——2′-5′A核。每个保护的单体接到载体上的收率是令人满意的,分别是100％,64％和89％。在合成过程中,我们对合成一系列中间体和制备载体的方法进行了改进,使产物收率大大提高,分离操作得以简化。     

Multiply-charged negative clusters of adenosine-5′-monophosphate in the gas phase

Multiply-charged noncovalent cluster anions of adenosine-5'-monophosphate (AMP) were formed by electrospray ionization (ESI). Ions in higher charge states were observed when the ions were accumulated in an ion trap with helium buffer gas before detection. We determined the smallest size (n(a)) or appearance size as a function of charge state (q), i.e., n(a) = 4 for q = 2, n(a) = 8 for q = 3, and n(a) = 13 for q = 4. The relation between n(a) and q can be described by a charged droplet model. When the size is larger than n(a) for a given q, the fragmentation pathway of an anion cluster is dominated by loss of neutral fragments. In contrast, when the size approaches the appearance size, only charged fragments are formed.     

A study of long range dipolar coupling constants in 4′-pentyloxy-4-biphenylcarbonitrile (5OCB)

It was shown previously that long range ²H-¹³ C dipolar coupling constants in liquid crystals could be obtained from the proton-decoupled ¹³C NMR spectrum of monodeuterated compounds. This approach has now been applied to the study of 4′-(1-deuteropentyloxy)-4-biphenylcarbonitrile (5OCB-d1). The 1¹³C spectrum of this compound gives nine ²H- ¹³C dipolar coupling constants for carbon ranging from one to six bonds away from the dueterium nucleus. The 2 H NMR and 5OCB-d1 gives the deuterium quadrupole coupling constant and the ¹H- ²H dipolar coupling constant for the α -CHD group. In addition, eleven 2-bond and two 3-bond ¹H- ¹³C dipolar coupling constants have been obtained for 4′-pentyloxy-4-biphenylcarbonitrile (non-deuterated 5OCB) by using the 2D ¹³C method of proton-encoded local field spectroscopy.