Nassim Usman

Bio: Nassim Usman is an academic researcher from Massachusetts Institute of Technology. The author has contributed to research in topics: Ribozyme & RNA. The author has an hindex of 35, co-authored 81 publications receiving 4693 citations. Previous affiliations of Nassim Usman include École Polytechnique Fédérale de Lausanne & McGill University.

Chemical synthesis of biologically active oligoribonucleotides using beta-cyanoethyl protected ribonucleoside phosphoramidites.

Abstract: The preparation of fully protected diisopropylamino-beta-cyanoethyl ribonucleoside phosphoramidites with regioisomeric purity greater than 99.95% is described. It is demonstrated that the combination of standard DNA protecting groups, 5'-O-DMT, N-Bz (Ade and Cyt), N-iBu (Gua), beta-cyanoethyl for phosphate, in conjunction with TBDMS for 2'-hydroxyl protection, constitutes a reliable method for the preparation of fully active RNA. Average stepwise coupling yields in excess of 99% were achieved with these synthons on standard DNA synthesizers. Two steps completely deprotect the oligoribonucleotide and workup is reduced to a fifteen minute procedure. Further, it is shown that the deprotected oligoribonucleotides are free from 5'-2' linkages. This methodology was applied to the chemical synthesis of a 24-mer microhelix, a 35-mer minihelix and two halves of a catalytic 'Hammerhead Ribozyme'. These oligoribonucleotides were directly compared in two distinct biochemical assays with enzymatically (T7 RNA polymerase) prepared oligoribonucleotides and shown to possess equal or better activity.

The molecular structure of the complex of Hoechst 33258 and the DNA dodecamer d(CGCGAATTCGCG)

Abstract: The crystal structure of the complex between the dodecamer d(CGCGAATTCGCG) and a synthetic dye molecule Hoechst 33258 was solved by X-ray diffraction analysis and refined to an R-factor of 15.7% at 2.25 A resolution. The crescent-shaped Hoechst compound is found to bind to the central four AATT base pairs in the narrow minor groove of the B-DNA double helix. The piperazine ring of the drug has its flat face almost parallel to the aromatic bisbenzimidazole ring and lies sideways in the minor groove. No evidence of disordered structure of the drug is seen in the complex. The binding of Hoechst to DNA is stabilized by a combination of hydrogen bonding, van der Waals interaction and electrostatic interactions. The binding preference for AT base pairs by the drug is the result of the close contact between the Hoechst molecule and the C2 hydrogen atoms of adenine. The nature of these contacts precludes the binding of the drug to G-C base pairs due to the presence of N2 amino groups of guanines. The present crystal structural information agrees well with the data obtained from chemical footprinting experiments.

RNA Hydration: A Detailed Look†,‡

Abstract: The crystal structure of the RNA duplex [r(CCCCGGGG)]2 has been refined to 1.46 A resolution with room temperature synchrotron diffraction data. This represents the highest resolution reported to date for an all-RNA oligonucleotide and is well beyond the best resolution ever achieved with an A-form DNA duplex. The analysis of the ordered hydration around the octamer duplex reveals conserved regular arrangements of water molecules in both grooves. In the major groove, all located first shell water molecules can be fitted into a pattern that is repeated through all eight base pairs, involves half the phosphate oxygens, and joins the two strands. In the minor groove, roughly across its narrowest dimension, tandem water molecules link the 2'-hydroxyl groups of adjacent nucleotides in base-pair steps in a similarly regular fashion. The structure provides evidence for an important role of the 2'-hydroxyl groups in the thermodynamic stabilization of RNA, beyond their known functions of locking the sugar pucker and mediating 3' --> 5' intrastrand O2'...O4' hydrogen bonds. The ribose 2'-hydroxyls lay the foundation for the enthalpic stability of the RNA relative to the DNA duplex, both as a scaffold for the water network in the minor groove and through their extensive individual hydration.

Water as an Active Constituent in Cell Biology

Abstract: When Szent-Gyorgyi called water the “matrix of life”,1 he was echoing an old sentiment. Paracelsus in the 16th century said that “water was the matrix of the world and of all its creatures.”2 But Paracelsus’s notion of a matrixsan active substance imbued with fecund, life-giving propertiess was quite different from the picture that, until very recently, molecular biologists have tended to hold of water’s role in the chemistry of life. Although acknowledging that liquid water has some unusual and important physical and chemical propertiessits potency as a solvent, its ability to form hydrogen bonds, its amphoteric naturesbiologists have regarded it essentially as the backdrop on which life’s molecular components are arrayed. It used to be common practice, for example, to perform computer simulations of biomolecules in a vacuum. Partly this was because the computational intensity of simulating a polypeptide chain was challenging even without accounting for solvent molecules too, but it also reflected the prevailing notion that water does little more than temper or moderate the basic physicochemical interactions responsible for molecular biology. What Gerstein and Levitt said 9 years ago remains true today: “When scientists publish models of biological molecules in journals, they usually draw their models in bright colors and place them against a plain, black background”.3 Curiously, this neglect of water as an active component of the cell went hand in hand with the assumption that life could not exist without it. That was basically an empirical conclusion derived from our experience of life on Earth: environments without liquid water cannot sustain life, and special strategies are needed to cope with situations in which, because of extremes of either heat or cold, the liquid is scarce.4-6 The recent confirmation that there is at least one world rich in organic molecules on which rivers and perhaps shallow seas or bogs are filled with nonaqueous fluidsthe liquid hydrocarbons of Titan7smight now bring some focus, even urgency, to the question of whether water is indeed a * E-mail: p.ball@nature.com. Philip Ball is a science writer and a consultant editor for Nature, where he worked as an editor for physical sciences for more than 10 years. He holds a Ph.D. in physics from the University of Bristol, where he worked on the statistical mechanics of phase transitions in the liquid state. His book H2O: A Biography of Water (Weidenfeld & Nicolson, 1999) was a survey of the current state of knowledge about the behavior of water in situations ranging from planetary geomorphology to cell biology. He frequently writes about aspects of water science for both the popular and the technical media.

Potent and persistent in vivo anti-HBV activity of chemically modified siRNAs

Abstract: The efficacy of lipid-encapsulated, chemically modified short interfering RNA (siRNA) targeted to hepatitis B virus (HBV) was examined in an in vivo mouse model of HBV replication. Stabilized siRNA targeted to the HBV RNA was incorporated into a specialized liposome to form a stable nucleic-acid-lipid particle (SNALP) and administered by intravenous injection into mice carrying replicating HBV. The improved efficacy of siRNA-SNALP compared to unformulated siRNA correlates with a longer half-life in plasma and liver. Three daily intravenous injections of 3 mg/kg/day reduced serum HBV DNA >1.0 log(10). The reduction in HBV DNA was specific, dose-dependent and lasted for up to 7 d after dosing. Furthermore, reductions were seen in serum HBV DNA for up to 6 weeks with weekly dosing. The advances demonstrated here, including persistence of in vivo activity, use of lower doses and reduced dosing frequency are important steps in making siRNA a clinically viable therapeutic approach.