2'-O-Methyloligoribonucleotides have been synthesised on solid phase from base protected 5'-O-dimethoxytrityl-2'-O-methylribonucleoside-3'-O-(2-cyanoethyl N,N-diisopropylphosphoramidites) using 5-(4-nitrophenyl)-1H-tetrazole as activator. Coupling yields greater than 99% were achieved, as judged by trityl cation release. The preparation of a modified 2'-deoxycytidine building block bearing an N4-(5-trifluoroacetylaminopentyl) spacer is also described. The latter compound enabled the chemical synthesis of 2'-O-methyloligoribonucleotide probes carrying several 5'- terminal biotinylation sites (in general four modified residues were used), which can be conveniently 32P end-labelled enzymatically using polynucleotide kinase. Used in conjunction with streptavidin-containing derivatives, such biotinylated probes have important applications in biochemical purification and electron microscopy of RNA-protein complexes. The 2'-O-methyloligoribonucleotides are completely resistant to degradation by either RNA or DNA specific nucleases. In contrast, nucleases with dual RNA/DNA specificity show a complete spectrum of cleavage rates.
alpha,gamma-Diketo acids (DKA) were discovered from screening as selective and reversible inhibitors of hepatitis C virus NS5b RNA-dependent RNA polymerase. The diketo acid moiety proved essential for activity, while substitution on the gamma position was necessary for selectivity and potency. Optimization led to the identification of a DKA inhibitor of NS5b polymerase with IC(50) = 45 nM, one of the most potent HCV NS5b polymerase inhibitors reported.
ABSTRACT2'-O-Methyl oligoribonucleotides have recently been introduced as antisense probes for studying RNA processing and for affinity purification of RNA-protein complexes. To identify RNA analogues with improved properties for antisense analysis, 2'--alkyl oligoribonucleotides were synthesized in which the alkyl moiety was either the threecarbon linear allyl group or the five-carbon branched 3,3-dimethylallyl group. Both these analogues were found to be completely resistant to degradation by either DNA-or RNAspecific nucleases. Use of biotinylated derivatives of the probes to affinity-select ribonucleoprotein particles from crude HeLa cell nuclear extracts showed that the presence of the bulky
Neuromedin U (NMU) is an endogenous peptide implicated in the regulation of feeding, energy homeostasis, and glycemic control, which is being considered for the therapy of obesity and diabetes. A key liability of NMU as a therapeutic is its very short half-life in vivo. We show here that conjugation of NMU to human serum albumin (HSA) yields a compound with long circulatory half-life, which maintains full potency at both the peripheral and central NMU receptors. Initial attempts to conjugate NMU via the prevalent strategy of reacting a maleimide derivative of the peptide with the free thiol of Cys34 of HSA met with limited success, because the resulting conjugate was unstable in vivo. Use of a haloacetyl derivative of the peptide led instead to the formation of a metabolically stable conjugate. HSA-NMU displayed long-lasting, potent anorectic, and glucose-normalizing activity. When compared side by side with a previously described PEG conjugate, HSA-NMU proved superior on a molar basis. Collectively, our results reinforce the notion that NMU-based therapeutics are promising candidates for the treatment of obesity and diabetes.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.