Expressão e purificação de domínios estruturais da proteína prion celular (PrPC) : ferramentas para a caracterização da interação entre PrPC e Stub1/Chip

Orientadora: Adriana Frohlich MercadanteMonografia (Bacharelado) - Universidade Federal do Paraná. Setor de Ciências Biológicas. Curso de Graduação em Ciências Biológica

Selective recognition of pyrimidine–pyrimidine DNA mismatches by distance-constrained macrocyclic bis-intercalators

Binding of three macrocyclic bis-intercalators, derivatives of acridine and naphthalene, and two acyclic model compounds to mismatch-containing and matched duplex oligodeoxynucleotides was analyzed by thermal denaturation experiments, electrospray ionization mass spectrometry studies (ESI-MS) and fluorescent intercalator displacement (FID) titrations. The macrocyclic bis-intercalators bind to duplexes containing mismatched thymine bases with high selectivity over the fully matched ones, whereas the acyclic model compounds are much less selective and strongly bind to the matched DNA. Moreover, the results from thermal denaturation experiments are in very good agreement with the binding affinities obtained by ESI-MS and FID measurements. The FID results also demonstrate that the macrocyclic naphthalene derivative BisNP preferentially binds to pyrimidine–pyrimidine mismatches compared to all other possible base mismatches. This ligand also efficiently competes with a DNA enzyme (M.TaqI) for binding to a duplex with a TT-mismatch, as shown by competitive fluorescence titrations. Altogether, our results demonstrate that macrocyclic distance-constrained bis-intercalators are efficient and selective mismatch-binding ligands that can interfere with mismatch-binding enzymes

Expressão e purificação de domínios estruturais da proteína prion celular (PrPC) : ferramentas para a caracterização da interação entre PrPC e Stub1/Chip

Orientadora: Adriana Frohlich MercadanteMonografia (Bacharelado) - Universidade Federal do Paraná. Setor de Ciências Biológicas. Curso de Graduação em Ciências Biológica

The Phenolic Antioxidant 3,5-dihydroxy-4-methoxybenzyl Alcohol (DHMBA) Prevents Enterocyte Cell Death under Oxygen-Dissolving Cold Conditions through Polyphyletic Antioxidant Actions

Cold preservation in University of Wisconsin (UW) solution is not enough to maintain the viability of the small intestine, due to the oxidative stress. The novel phenolic antioxidant 3,5-dihydroxy-4-methoxybenzyl alcohol (DHMBA) has dual properties to reduce oxidative stress, radical scavenging, and antioxidant protein induction, in other cells. This study was designed to determine whether DHMBA reduces cold preservation injury of enterocytes, and to identify the effector site. Enterocytes were subjected to 48-h cold preservation under atmosphere in UW solution (+/- DHMBA), and then returned to normal culture to replicate reperfusion of the small intestine after cold preservation. At the end of cold preservation (ECP) and at 1, 3, 6, and 72 h after rewarming (R1h, R3h, R6h, and R72h), we evaluated cell function and the injury mechanism. The results showed that DHMBA protected mitochondrial function mainly during cold preservation, and suppressed cell death after rewarming, as shown by the MTT, ATP, mitochondrial membrane potential, LDH, and lipid peroxidation assays, together with enhanced survival signals (PI3K, Akt, p70S6K) and induction of antioxidant proteins (HO-1, NQO-1, TRX-1). We found that DHMBA mitigates the cold-induced injury of enterocytes by protecting the mitochondria through direct and indirect antioxidative activities