Current Chemical Biology - Volume 1, Issue 3, 2007

DNA polymerases catalyze the addition of mononucleotides into a growing polymer using a DNA template as a guide for directing each incorporation event. The efficiency and fidelity of this biological process have been historically attributed to the ability of the DNA polymerase to coordinate proper hydrogen-bonding interactions between the incoming nucleotide with the templating nucleobase. However, the strength of this model has been weakened since several laboratories have demonstrated that non-natural nucleotides, i.e., those devoid of typical hydrogen-bonding capabilities, can be utilized by DNA polymerases with varying degrees of efficiencies. This review provides a comprehensive summary of current research efforts leading to the development and implementation of these analogs as probes for DNA polymerase function and activity. The ability of various non-natural purines and pyrimidines to be incorporated opposite templating nucleobases suggests that polymerization efficiency is not directly influenced by hydrogen-bonding interactions but rather by the overall shape and size of the formed base-pair. Conflicting evidence is obtained when the dynamics of nucleotide incorporation is assessed using nucleic acid containing permutations in hydrogen bonding capabilities or completely devoid of these interactions. With respect to replication opposite an abasic site, it appears that the π-electron surface area and desolvation properties of the incoming nucleotide play a significant role for facilitating incorporation. This information has lead to the development of new models for DNA polymerization as well as toward strategies for novel biotechnology platforms and unique chemotherapeutic agents.

Carbohydrates are often referred to as the third molecular chain of life, and they represent the most important post-genomics research targets. Recent reports of their biological functions in signal transduction and cellular recognition have made them promising pharmaceutical targets and disease markers. Like nucleic acids and proteins, the three-dimensional (3D) structures of carbohydrates are important for their molecular functions. Currently, the most abundant source of biological carbohydrate structures is the Protein Data Bank (PDB). As its name indicates, the PDB is a protein structure database. However, carbohydrate structures have often been determined in complexes with protein molecules, as enzyme substrates, lectin ligands, or post-translational modifications. As of Jul 2006, the PDB contained 6,421 carbohydrate-protein complex structures, and the number is increasing rapidly. In this review, the current status of the PDB as the carbohydrate structure database, and the features of several databases derived from the PDB will be summarized. We will also introduce an overview of the bioinformatics tools currently available for analyses of carbohydrate 3D structures.

The accuracy of the docking of a ligand in its modeled binding-site depends on the reliability of this model. To provide a model with experimental support, we have developed an engineered affinity labeling method combining cysteine-reactive probes with substitutions of putative site-lining residues into cysteines. This strategy amounts to building chemical sensors for the proximity of the substituted cysteines; it requires an activity or binding assay to monitor the irreversible occupancy of the site by the reactive ligand. Using affinity probes made reactive in different positions, the docking of the ligand can be inferred from the observed pattern of coupling reactions. The method involves three steps: ligand chemistry, mutagenesis and biological assays, which are detailed and scrutinized in the review: lead selection, ligand derivatization, and evaluation of the affinity probes (stability, reactivity and biological properties) for the ligands; positional selection and mutant properties for the cysteine substitutions; functional controls and assays for the analysis of the irreversible reactions. Examples illustrate the different criteria of concern; the data are interpreted in terms of binding-site structure and function. Potentially, the method can explore protein dynamics, since its targets are full-length membrane-inserted heteromeric proteins: it can detect subtype-dependent or activation-induced conformational changes.

The history of inorganic pharmacology can be traced to antiquity with the medicinal use of inorganic salts, coordination and organometallic compounds. The clinical applications of metal-based drugs today are limited, but extremely significant. The most common metallo-therapeutic drugs are platinum, gold and bismuth compounds used in anticancer protocols and in the treatment of rheumatoid arthritis and gastric and duodenal ulcers, respectively. Platinum(II)-derivatives are the most widely prescribed anticancer agents, especially for polychemotherapy. Years of clinical experience have yielded detailed information about the quantitative structure-activity relationship (QSAR), pharmacokinetics and mechanisms of action of Pt-drugs. The accuracy of this information depends on precise measurement of Pt levels in body fluids, tissues, cells and DNA. Inductively Coupled Plasma - Mass Spectrometry (ICP-MS) offers higher sensitivity and accuracy than conventional analytical techniques, making it possible to detect trace concentrations of Pt-drugs at truly pharmacological concentrations. Increased knowledge about the action and fate of Pt-drugs may lead to important insights for the development of new metallo-pharmaceuticals with even wider applications.

This review briefly describes three unique types of solid surfaces, i.e., honeycomb polymer films, thermo-responsive polymergrafted glass surfaces and super water-repellent surfaces with a fractal structure. Details of preparation and physical properties are presented first followed by highlights of recent progress in cell culture on the solid surfaces.

Over 20 million patients in the United States alone each year receive a general anesthetic for a surgical procedure. Nevertheless, molecular mechanisms of volatile general anesthetic action remain poorly understood. The favored sites of action in the central nervous system are currently a variety of plasma membrane proteins including the Cys-loop superfamily of ligand-gated ion channels and the N-methyl-D-aspartate receptor, because volatile general anesthetics are able to alter the ion conducting properties of these proteins. Volatile general anesthetics are only capable of relatively weak interactions with macromolecular targets, precluding the use of conventional radioligand binding assays for identifying central nervous system targets. In order to overcome this significant technical obstacle, other approaches to monitor volatile general anesthetic binding have been developed that rely on 19F-nuclear magnetic resonance spectroscopy, photoaffinity labeling with halothane, fluorescence spectroscopy, and isothermal titration calorimetry. These techniques have allowed the determination of volatile general anesthetic dissociation constants for a number of different protein complexes. The effect of a bound volatile general anesthetic on protein stability, flexibility, and overall structure has been investigated in recent years, and the results suggest fundamental mechanisms whereby these important clinical compounds reversibly alter protein function.

Reactive oxygen species (ROS) are oxygen-containing molecules that possess unpaired electrons. ROS are a normal component of cellular life, and accumulating evidence suggests that these molecules play critical roles in many important signal transduction pathways. However, maintaining a balance between ROS production and elimination is a crucial component of cellular homeostasis. Unregulated increases in cellular ROS can inflict significant physical damage on subcellular structures, such as mitochondria and lipid membranes. Furthermore, accumulating evidence asserts that ROS can play an aberrant role in cellular signaling if their production is left unchecked. This review is a discussion of the interaction between ROS and the Janus Kinase 2 (Jak2) signal transduction pathway in the context of three highly prevalent diseases; diabetes, atherosclerosis and cardiac ischemia-reperfusion injury. ROS-mediated Jak2 activation contributes to the progression of diabetic nephropathy, atherosclerosis and acute cardiac ischemia-reperfusion injury. Interestingly, this mechanism also appears to play a cardioprotective role in preconditioned cardiac ischemia-reperfusion injury. Currently, its role in diabetic cardiomyopathy is unclear. Thus, the Jak2/ROS relationship appears to have significant consequences for human health, as indicated by its prominent role in several highly prevalent disorders.

Since the description of the synthetic chemical clofibrate in 1962, various derivatives of fibrates with a diversity of chemical structures have been developed. Several of these are used clinically to treat dyslipidemia because they are generally effective in lowering elevated plasma triglycerides and cholesterol. Studies suggest that several biochemical mechanisms underlie fibrate-mediated modulation of lipoprotein and related metabolites. These mechanisms are: 1) induced lipoprotein lipolysis; 2) induced hepatic fatty acid uptake and reduced hepatic triglyceride formation; 3) amplified removal of low density lipoprotein (LDL) particles; 4) reduced neutral lipid (cholesteryl ester and triglyceride) exchange between very low density lipoprotein (VLDL) and high density lipoprotein (HDL) resulting from decreased plasma levels of triglyceride-rich lipoprotein (TRL); and 5) increased HDL production and stimulation of reverse cholesterol transport. Recent studies of structure-based inhibitor design strategy revealed that an independent enzyme, aldose reductase (AR), is a target of fibrate activity, an additional biochemical mechanism. AR has been implicated as a major player in the development of diabetes and diabetic complications because of its ability to catalyze the conversion of glucose to sorbitol. This article discusses various targets of fibrate action, biochemical pathways and commonalities in potential molecular interactions.

This review article deals with different aspects of the dioxaspiro[2.4]heptanes, such as their presence in Nature, biogenetic origin, biological activity, as well as the different strategies for their synthesis and their chemical reactivity. In addition, selected total syntheses of natural products containing this moiety is presented, paying especial attention to the steps involving the generation of the dioxaspiro[ 2.4]heptane unit.