Molecular Beacon DNA Probes with Fluorescein Bifluorophore

Russian Journal of Bioorganic Chemistry - An azido-derivative of a fluorescein bifluorophore was obtained and used for the synthesis of “molecular beacon”-type oligonucleotide...     

Reagents For The Selective Immobilization Of Oligonucleotides On Solid Supports

New reagents (CPGs and phosphoramidites) for automatic solid phase synthesis of modified oligonucleotides were designed. Three oligonucleotides carrying fluorescent label at the 5′-terminus and an anchor group at the 3′-terminus were prepared and their immobilization in orthogonal conditions on solid supports was studied.     

Isotope-reinforced polyunsaturated fatty acids protect yeast cells from oxidative stress

The facile abstraction of bis-allylic hydrogens from polyunsaturated fatty acids (PUFAs) is the hallmark chemistry responsible for initiation and propagation of autoxidation reactions. The products of these autoxidation reactions can form cross-links to other membrane components and damage proteins and nucleic acids. We report that PUFAs deuterated at bis-allylic sites are much more resistant to autoxidation reactions, because of the isotope effect. This is shown using coenzyme Q-deficient Saccharomyces cerevisiae coq mutants with defects in the biosynthesis of coenzyme Q (Q). Q functions in respiratory energy metabolism and also functions as a lipid-soluble antioxidant. Yeast coq mutants incubated in the presence of the PUFA α-linolenic or linoleic acid exhibit 99% loss of colony formation after 4 h, demonstrating a profound loss of viability. In contrast, coq mutants treated with monounsaturated oleic acid or with one of the deuterated PUFAs, 11,11-D₂-linoleic or 11,11,14,14-D₄-α-linolenic acid, retain viability similar to wild-type yeast. Deuterated PUFAs also confer protection to wild-type yeast subjected to heat stress. These results indicate that isotope-reinforced PUFAs are stabilized compared to standard PUFAs, and they protect coq mutants and wild-type yeast cells against the toxic effects of lipid autoxidation products. These findings suggest new approaches to controlling ROS-inflicted cellular damage and oxidative stress.     

Control of lysyl oxidase activity through site-specific deuteration of lysine

Lysyl oxidase (LOX) is implicated in several extracellular matrix related disorders, including fibrosis and cancer. Methods of inhibition of LOX in vivo include antibodies, copper sequestration and toxic small molecules such as β-aminopropionitrile. Here, we propose a novel approach to modulation of LOX activity based on the kinetic isotope effect (KIE). We show that 6,6-d₂-lysine is oxidised by LOX at substantially lower rate, with apparent deuterium effect on V_max/K_m as high as 4.35 ± 0.22. Lys is an essential nutrient, so dietary ingestion of D₂Lys and its incorporation via normal Lys turnover suggests new approaches to mitigating LOX-associated pathologies.     

Small amounts of isotope-reinforced polyunsaturated fatty acids suppress lipid autoxidation

Polyunsaturated fatty acids (PUFAs) undergo autoxidation and generate reactive carbonyl compounds that are toxic to cells and associated with apoptotic cell death, age-related neurodegenerative diseases, and atherosclerosis. PUFA autoxidation is initiated by the abstraction of bis-allylic hydrogen atoms. Replacement of the bis-allylic hydrogen atoms with deuterium atoms (termed site-specific isotope-reinforcement) arrests PUFA autoxidation due to the isotope effect. Kinetic competition experiments show that the kinetic isotope effect for the propagation rate constant of Lin autoxidation compared to that of 11,11-D(2)-Lin is 12.8 ± 0.6. We investigate the effects of different isotope-reinforced PUFAs and natural PUFAs on the viability of coenzyme Q-deficient Saccharomyces cerevisiae coq mutants and wild-type yeast subjected to copper stress. Cells treated with a C11-BODIPY fluorescent probe to monitor lipid oxidation products show that lipid peroxidation precedes the loss of viability due to H-PUFA toxicity. We show that replacement of just one bis-allylic hydrogen atom with deuterium is sufficient to arrest lipid autoxidation. In contrast, PUFAs reinforced with two deuterium atoms at mono-allylic sites remain susceptible to autoxidation. Surprisingly, yeast treated with a mixture of approximately 20%:80% isotope-reinforced D-PUFA:natural H-PUFA are protected from lipid autoxidation-mediated cell killing. The findings reported here show that inclusion of only a small fraction of PUFAs deuterated at the bis-allylic sites is sufficient to profoundly inhibit the chain reaction of nondeuterated PUFAs in yeast.     

Reagents for the selective immobilization of oligonucleotides on solid supports

New reagents (CPGs and phosphoramidites) for automatic solid phase synthesis of modified oligonucleotides were designed. Three oligonucleotides carrying fluorescent label at the 5'-terminus and an anchor group at the 3'-terminus were prepared and their immobilization in orthogonal conditions on solid supports was studied.     

5-arylethynyl-2'-deoxyuridines: energy transfer and SNP-detection

Energy transfer between different fluorescent 5-alkynyl-2' -deoxyuridines in complementary and mismatched duplexes was studied.     

Prediction of the three-dimensional structure of aflatoxin of Aspergillus flavus by homology modelling

Aflatoxins are polyketide-derived secondary metabolites produced by Aspergillus spp. The toxic effects of aflatoxins have adverse consequences for human health and agricultural economics. The aflR gene, a regulatory gene for aflatoxin biosynthesis, encodes a protein containing a zinc-finger DNA-binding motif. AFLR-Protein three-dimensional model was generated using Robetta server. The modeled AFLR-Protein was further optimization and validation using Rampage. In the simulations, we monitored the backbone atoms and the C-α-helix of the modeled protein. The low RMSD and the simulation time indicate that, as expected, the 3D structural model of AFLR-protein represents a stable folding conformation. This study paves the way for generating computer molecular models for proteins whose crystal structures are not available and which would aid in detailed molecular mechanism of inhibition of aflatoxin.     

Computer Modeling and Synthesis of Potential Inhibitors of Tyrosine Kinase BCR-ABL with the T315I Mutation

Russian Journal of Bioorganic Chemistry - A comparative analysis of the interaction of the chimeric protein BCR-ABL, of the normal type and with the T315I mutation, with known inhibitors as well as...