RESUMO
Lipids in Escherichia coli and Bacillus subtilis were analyzed by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry and TOF/TOF tandem mass spectrometry. Lipids were extracted from bacterial cells using an equal volume mixture of dichloromethane, ethanol, and water, which formed a biphasic system with the lipids in the organic layer. The resulting mass spectra of the extracts from both bacteria showed a series of peaks corresponding to sodiated phospholipids - primarily phosphatidylethanolamines (PE) and phosphatidylglycerols (PG). The relative amounts of the phospholipids and the fatty acid compositions inferred from the spectra were in good agreement with previously reported values from GC/MS and thin-layer chromatography studies. E. coli and B. subtilis were easily differentiated by dissimilarities in the composition and relative amounts of the phospholipids present as well as by the presence of lysyl-phosphatidylglycerol and diglucosyl diglycerides solely in the B. subtilis mass spectra. Changes in lipid content in the bacteria during their growth phases were also monitored. In E. coli, the spectra indicated an increase in the amount of the unique C(cy-17) fatty acid (in which the fatty acid chain contains a cyclopropane ring) formed during exponential growth. During stationary growth, the spectra indicated an increase in the amount of saturated fatty acids. In B. subtilis, the phospholipid composition remained relatively unchanged during exponential growth, but the amount of PG slightly decreased while the amount of PE slightly increased during stationary growth. No significant changes were observed for the lysyl-phosphatidylglycerols or glycolipids during the exponential or stationary growth phases.
RESUMO
The glutathione S-transferase Mu class (GSTM) genes encode phase II metabolism enzymes that are involved in the detoxification of various carcinogens and drugs. Some genetic polymorphisms in GSTM genes are related to disease phenotypes and drug-metabolism differences in the population. Polymorphisms that alter gene-splicing patterns are functionally very important because they often lead to the insertion or deletion of many amino acids. To identify inter-individual differences in the splicing pattern of the GSTM4 gene, we used reverse transcriptase polymerase chain reaction (RT-PCR) to screen cDNA from 96 human liver samples. We discovered a novel splice variant of GSTM4 that resulted from tandem skipping of exons 4 and 5. This exon-skipping event is associated with a mutation at the splice acceptor site in intron 4.
Assuntos
Éxons , Glutationa Transferase/genética , Mutação Puntual , Splicing de RNA , DNA Complementar/genética , DNA Complementar/metabolismo , DNA Recombinante/metabolismo , Variação Genética , Glutationa Transferase/química , Glutationa Transferase/metabolismo , Humanos , Íntrons , Fígado/enzimologia , Fígado/metabolismo , Modelos Genéticos , Polimorfismo Genético , Conformação Proteica , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
The spread of H5N1 avian influenza viruses (AIVs) from China to Europe has raised global concern about their potential to infect humans and cause a pandemic. In spite of their substantial threat to human health, remarkably little AIV whole-genome information is available. We report here a preliminary analysis of the first large-scale sequencing of AIVs, including 2196 AIV genes and 169 complete genomes. We combine this new information with public AIV data to identify new gene alleles, persistent genotypes, compensatory mutations, and a potential virulence determinant.
Assuntos
Genes Virais , Virus da Influenza A Subtipo H5N1/genética , Vírus da Influenza A/genética , Proteínas não Estruturais Virais/química , Fatores de Virulência/química , Animais , Aves/virologia , Biologia Computacional , Genoma Viral , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H2N2/genética , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza A Subtipo H3N8/genética , Virus da Influenza A Subtipo H5N1/química , Virus da Influenza A Subtipo H5N1/patogenicidade , Vírus da Influenza A Subtipo H5N2/genética , Vírus da Influenza A Subtipo H7N7/genética , Vírus da Influenza A Subtipo H9N2/genética , Vírus da Influenza A/química , Vírus da Influenza A/isolamento & purificação , Vírus da Influenza A/patogenicidade , Influenza Aviária/virologia , Influenza Humana/virologia , Dados de Sequência Molecular , Mutação , Filogenia , RNA Viral/genética , Vírus Reordenados/genética , Análise de Sequência de DNA , Proteínas não Estruturais Virais/genética , Proteínas Virais/química , Proteínas Virais/genética , Fatores de Virulência/genéticaRESUMO
To examine the possibility that inter-individual differences in splicing partially explain the observed differences in CYP2D6 activity, we amplified its full-length cDNA in 96 human liver RNA samples and discovered five splice variants: intron 5 retention, intron 6 retention, intron 5 and intron 6 double retention, exon 3 skipping, and partial intron 1 retention. All of the CYP2D6 splice variants we identified are probably nonfunctional transcripts. Substantial inter-individual variation in the proportions of the CYP2D6 transcript represented by splice variants, measured by real-time PCR, suggests that the presence of these splice variants contributes to the population variation in CYP2D6 activity. Relatively high levels of intron 6 retention were not correlated with the newly discovered single nucleotide polymorphism 2988G > A in intron 6 (CYP2D6*41) but did correlate with the more common CYP2D6*34 allele. Our study prompts further investigations to explore the effect of these splice variants on drug metabolism.