基于结构的O-GlcNAc转移酶抑制剂的虚拟筛选

利用O-GlcNAc 转移酶同UDP-GlcNAc复合物的晶体结构,针对其催化位点,对ZINC库中的7134792个分子和FOG库中的4287550个分子进行三轮(HTVS、SP、XP)虚拟筛选,结果发现具有更好类药性的FOG库中包含更多对接得分更低的小分子,且具有更多新颖的化学片段.ZINC库中具有较低对接得分的分子可分为2类,分别占据UDP-GlcNAc的UDP和GlcNAc的结合位置,在此基础上设计得到的分子具有更好的对接得分.证明FOG分子库具有产生更多对接得分更低的分子,所预测和设计的小分子化合物可以成为潜在的抑制剂药物分子.     

Virtual Screening of Human O-GlcNAc Transferase Inhibitors

O-GlcNAc transferase (OGT) is one of essential mammalian enzymes, which catalyze the transfer of N-acetylglucosamine from UDP-N-acetylglucosamine (UDP-GlcNAc) to hydroxyl groups of serines and threonines (Ser/Thr) in proteins. Dysregulations of cellular O-GlcNAc have been implicated in diabetes, neurodegenerative disease, and cancer, which brings great interest in developing potent and speci c small-molecular OGT inhibitors. In this work, we performed virtual screening on OGT catalytic site to identify potential inhibitors. 7134792 drug-like compounds from ZINC (a free database of commercially available compounds for virtual screening) and 4287550 compounds generated by FOG (fragment optimized growth program) were screened and the top 116 compounds ranked by docking score were analyzed. By comparing the screening results, we found FOG program can generate more compounds with better docking scores than ZINC. The top ZINC compounds ranked by docking score were grouped into two classes, which held the binding positions of UDP and GlcNAc of UDPGlcNAc. Combined with individual fragments in binding pocket, de novo compounds were designed and proved to have better docking score. The screened and designed compounds may become a starting point for developing new drugs.     

Butylacrylate‐nucleobase Conjugates as Targets for Two‐step Redox Labeling of DNA with an Osmium Tetroxide Complex

Modification of nucleic acids with osmium tetroxide reagents (Os,L, such as OsO₄,2,2′‐bipyridine, Os,bpy) has been applied in redox DNA labeling, in probing DNA structure as well as in studies of DNA interactions with other molecules. In natural DNA, primarily thymine residues form adducts with the Os,bpy in a structure selective manner. In this paper we introduce a new two‐step technique of DNA modification with the electroactive Os,bpy, consisting in enzymatic construction of DNA bearing butyl acrylate (BA) moieties attached to uracil at C5 or to 7‐deaza adenine at C7, followed by chemical modification of a reactive C=C double bond in the acrylate residue. We demonstrate a facile modification of the BA conjugates in both single‐ and double‐stranded (ds) DNA under conditions when modification within the nucleobase rings in ds DNA is hindered. Various DNA−Os,bpy adducts can easily be analyzed electrochemically and distinguished by different redox potentials. The two‐step procedure appears to be applicable in osmium redox labelling of ds DNA.     

NMR structure of the peptidyl transferase RNA inhibitor antibiotic amicetin

In this communication, we report the solution state NMR structure determination of the peptidyl transferase RNA inhibitor antibiotic amicetin. We have successfully characterised the NMR spectrum of amicetin using a range of homo- and heteronuclear NMR techniques. Using experimental ROE-based distance and 1H--1H scalar coupling derived dihedral angle geometrical constraints as input into the three-dimensional structure determination protocol, we have generated an energy-minimised average structure of the antibiotic. Amicetin adopts a stable well-folded conformation in solution, mediated by a network of hydrogen bonds caused by proton donor and acceptor groups at either end of the molecule. The NMR structure of amicetin shows that the cytosine moiety occupies the critical turn position within the fold, which may be structurally significant for interaction with peptidyl transferase ribosomal RNA. The structure is distinctly different from the published X-ray crystal structure of amicetin in which it adopts a linear, extended chain-like conformation with a number of intermolecular hydrogen bonds. In addition to structure, we have probed the dynamics of amicetin in solution and have observed retarded exchange of the amide proton involved in folding. We have also characterised the ionisation properties of amicetin by carrying out NMR pH titration and measuring the pKa of the primary and tertiary amino groups, 7.27 and 7.52, respectively, which are in agreement with the reported values in literature. Solving the NMR structure of amicetin provides a valuable opportunity to determine the structure of its complex with RNA in solution state.     

An Inhibitor of Glutathione S-Transferase Omega 1 that Selectively Targets Apoptotic Cells

On (cell) suicide watch: The increased permeability of apoptotic cells was used to identify a peptide-based, covalent inhibitor (NJP2) for glutathione S-transferase omega (GSTO1) that is highly selective for apoptotic cells, with no inhibition of healthy cells. This apoptosis-specific peptide could also be used for imaging programmed cell death.     

Label‐free Voltammetric Detection of Products of Terminal Deoxynucleotidyl Transferase Tailing Reaction

A label‐free approach that takes advantage of intrinsic electrochemical activity of nucleobases has been applied to study the products of terminal deoxynucleotidyl transferase (TdT) tailing reaction. DNA homooligonucleotides A₃₀, C₃₀ and T₃₀ were used as primers for the tailing reaction to which a dNTP – or a mixture of dNTPs – and TdT were added to form the tails. Electrochemical detection enabled study of the tailing reaction products created by various combinations of primers and dNTPs, with pyrolytic graphite electrode (PGE) being suitable for remarkably precise analysis of the length of tailing reaction products. Furthermore, the hanging mercury drop electrode (HMDE) was able to reveal formation of various DNA structures, such as DNA hairpins and G‐quadruplexes, which influence the behavior of DNA molecules at the negatively charged surface of HMDE. Thus, the described approach proves to be an excellent tool for studying the TdT tailing reactions and for exploring how various DNA structures affect both the tailing reactions and electrochemical behavior of DNA oligonucleotides at electrode surfaces.     

Monitoring tissue inflammation and responses to drug treatments in early stages of mice bone fracture using 50 MHz ultrasound

Bone fracture induces moderate inflammatory responses that are regulated by cyclooxygenase-2 (COX-2) or 5-lipoxygenase (5-LO) for initiating tissue repair and bone formation. Only a handful of non-invasive techniques focus on monitoring acute inflammation of injured bone currently exists. In the current study, we monitored in vivo inflammation levels during the initial 2 weeks of the inflammatory stage after mouse bone fracture utilizing 50 MHz ultrasound. The acquired ultrasonic images were correlated well with histological examinations. After the bone fracture in the tibia, dynamic changes in the soft tissue at the medial-posterior compartment near the fracture site were monitored by ultrasound on the days of 0, 2, 4, 7, and 14. The corresponding echogenicity increased on the 2nd, 4th, and 7th day, and subsequently declined to basal levels after the 14th day. An increase of cell death was identified by the positive staining of deoxynucleotidyl transferase dUTP nick end-labeling (TUNEL) assay and was consistent with ultrasound measurements. The increases of both COX-2 and Leukotriene B4 receptor 1 (BLT₁, 5-LO-relative receptor), which are regulators for tissue inflammation, in the immunohistochemistry staining revealed their involvement in bone fracture injury. Monitoring the inflammatory response to various non-steroidal anti-inflammatory drugs (NSAIDs) treatments was investigated by treating injured mice with a daily oral intake of aspirin (Asp), indomethacin (IND), and a selective COX-2 inhibitor (SC-236). The Asp treatment significantly reduced fracture-increased echogenicity (hyperechogenicity, p < 0.05) in ultrasound images as well as inhibited cell death, and expression of COX-2 and BLT₁. In contrast, treatment with IND or SC-236 did not reduce the hyperechogenicity, as confirmed by cell death (TUNEL) and expression levels of COX-2 or BLT₁. Taken together, the current study reports the feasibility of a non-invasive ultrasound method capable of monitoring post-fracture tissue inflammation that positively correlates with histological findings. Results of this study also suggest that this approach may be further applied to elucidate the underlying mechanisms of inflammatory processes and to develop therapeutic strategies for facilitating fracture healing.     

Total Synthesis of Carpacin and its Geometric Isomer as a Cancer Chemopreventer

Carpacin ( 1a ), an antidepressant in Asiatic folk medicine from the Carpano tree, is achieved in which the longest linear sequence is only four steps in over all yield 67% from commercially available Sesamol. The key transformations in the synthesis are the selective palladium‐catalyzed coupling reactions of aryl bromide with Grignard reagents. The first preparation of its geometric isomer ( 1b ) is described. Highlights of the synthesis include Pd‐catalyzed coupling, selective hydrogenation, and Wittig reactions. Carpacin was examined as a potential inhibitor of carcinogenesis.     

Silicon nanowires as field-effect transducers for biosensor development: A review

The unique electronic properties and miniaturized dimensions of silicon nanowires (SiNWs) are attractive for label-free, real-time and sensitive detection of biomolecules. Sensors based on SiNWs operate as field effect transistors (FETs) and can be fabricated either by top–down or bottom–up approaches. Advances in fabrication methods have allowed for the control of physicochemical and electronic properties of SiNWs, providing opportunity for interfacing of SiNW-FET probes with intracellular environments. The Debye screening length is an important consideration that determines the performance and detection limits of SiNW-FET sensors, especially at physiologically relevant conditions of ionic strength (>100 mM). In this review, we discuss the construction and application of SiNW-FET sensors for detection of ions, nucleic acids and protein markers. Advantages and disadvantages of the top–down and bottom–up approaches for synthesis of SiNWs are discussed. An overview of various methods for surface functionalization of SiNWs for immobilization of selective chemistry is provided in the context of impact on the analytical performance of SiNW-FET sensors. In addition to in vitro examples, an overview of the progress of use of SiNW-FET sensors for ex vivo studies is also presented. This review concludes with a discussion of the future prospects of SiNW-FET sensors.