RESUMO
RNA contains diverse modifications that exert an important influence in a variety of cellular processes. So far, more than 150â modifications have been identified in various RNA species, mainly in ribosomal RNA (rRNA), transfer RNA (tRNA), and messenger RNA (mRNA). In contrast to rRNA, tRNA, and mRNA, the known modifications in small RNA species have been primarily limited to 2'-O-ribose methylation in plants and inosine in mammals. The methylation of small RNAs in mammals is still unclear. Current methods widely used in the characterization of small RNAs are mainly based on the strategy of nucleic acid hybridization and sequencing, which cannot characterize modifications in small RNAs. Herein, we have systematically investigated modifications in small RNAs composed of 16-28â nucleotides (nt) by establishing an effective isolation and neutral enzymatic digestion of small RNAs in combination with liquid chromatography/electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). This method allowed us to simultaneously detect 57â different types of nucleoside modification. By using this approach, we revealed 24â modifications in small RNAs comprising 16-28â nt from human cells. In addition, we found that the obesity-associated protein (FTO) may demethylate N6 -methyladenosine (m6 A) and N6 ,2'-O-dimethyladenosine (m6 Am) in small RNAs of 16-28â nt. Our study demonstrates the existence of diverse modifications in small RNAs composed of 16-28â nt, which may promote in-depth understanding of the regulatory roles of noncoding RNAs.
Assuntos
Adenosina/análogos & derivados , Dioxigenase FTO Dependente de alfa-Cetoglutarato/metabolismo , Nucleosídeos/metabolismo , Nucleotídeos/metabolismo , RNA Mensageiro/metabolismo , RNA de Transferência/metabolismo , RNA/química , Adenosina/química , Dioxigenase FTO Dependente de alfa-Cetoglutarato/química , Animais , Cromatografia Líquida , Humanos , Metilação , Nucleosídeos/química , Nucleotídeos/química , RNA Mensageiro/química , RNA de Transferência/química , Espectrometria de Massas em TandemRESUMO
A series of dicationic porphyrins were found to specifically recognize loop structures of oligodeoxynucleotides, and to selectively oxidize guanine residues upon photo-irradiation at micromolar concentrations. These compounds could, thus, be employed as promising structural probes for specific secondary DNA structures. The production of singlet oxygen ((1)O2) is responsible for the DNA-modification profiles. Both UV-titration and thermal-melting experiments indicate that the strong affinity of the charged porphyrins towards DNA loops is responsible for these molecular-recognition phenomena.
Assuntos
DNA/genética , DNA/metabolismo , Porfirinas/genética , Porfirinas/metabolismo , Sítios de Ligação/fisiologia , DNA/química , Conformação de Ácido Nucleico , Espectrofotometria Ultravioleta/métodosRESUMO
A series of biphenol-derived quaternary ammonium salts, originally developed as DNA-cross-linking agents, and carrying either two (i.e., 1) or four (i.e., 2) net positive charges, were investigated for their in vitro DNA-transcription- and acetylcholinesterase (AChE)-inhibitory activities. The effects of charge and type of linker between the two phenolic residues were systematically investigated. Several compounds showed good activities in both tests, which makes them potential lead candidates for drug design.
Assuntos
Acetilcolinesterase/metabolismo , Inibidores da Colinesterase/farmacologia , Fenóis/química , Compostos de Amônio Quaternário/farmacologia , Inibidores da Colinesterase/química , Dimerização , Desenho de Fármacos , Compostos de Amônio Quaternário/químicaRESUMO
The interaction between CT-DNA and the zinc phthalocyanine ZnPc (1) was studied by UV/VIS and fluorescence titration, as well as by thermal denaturation. ZnPc was found to strongly bind to CT-DNA (K(app)=7.35 x 10(5) M(-1)) in a non-intercalative mode. The photosensitized cleavage of pBR322 DNA was found to efficiently proceed via singlet-oxygen ((1)O(2)) production. Further, ZnPc (1) caused site-specific scission of guanine (G) bases around the bulge of the hairpin oligonucleotides OD1-OD3, as clearly shown by gel-electrophoresis experiments.