RESUMO
A large number of halogenated compounds is produced by chemical synthesis. Some of these compounds are very toxic and cause enormous problems to human health and to the environment. Investigations on the degradation of halocompounds by microorganisms have led to the detection of various dehalogenating enzymes catalyzing the removal of halogen atoms under aerobic and anaerobic conditions involving different mechanisms. On the other hand, more than 3500 halocompounds are known to be produced biologically, some of them in great amounts. Until 1997, only haloperoxidases were thought to be responsible for incorporation of halogen atoms into organic compounds. However, recent investigations into the biosynthesis of halogenated metabolites by bacteria have shown that a novel type of halogenating enzymes, FADH(2)-dependent halogenases, are involved in biosyntheses of halogenated metabolites. In every gene cluster coding for the biosynthesis of a halogenated metabolite, isolated so far, one or several genes for FADH(2)-dependent halogenases have been identified.
Assuntos
Halogênios/metabolismo , Hidrocarbonetos Halogenados/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH , Sequência de Aminoácidos , Bactérias/metabolismo , Biodegradação Ambiental , Catálise , Sequência Conservada , Enzimas/genética , Enzimas/metabolismo , Ácidos Graxos Dessaturases , Flavina-Adenina Dinucleotídeo/análogos & derivados , Flavina-Adenina Dinucleotídeo/metabolismo , Halogênios/química , Hidrocarbonetos Halogenados/química , Oxirredução , Alinhamento de SequênciaRESUMO
BACKGROUND: This study describes the evaluation of the expression patterns of prostate-related transcripts in 106 matched prostate tissues from prostatectomies as predictors for prostate cancer (PCa). METHODS: Quantitative PCR (QPCR) assays with site-specific hybridization probes were established for four housekeeping genes (GAPDH, HPRT, PBGD, TBP) and nine prostate-related genes (AibZIP, D-GPCR, EZH2, PCA3, PDEF, prostein, PSA, PSCA, TRPM8). RESULTS: The relative mRNA expression levels of AibZIP, D-GPCR, EZH2, PCA3, PDEF, PSA, TRPM8 (all P < 0.001) and prostein (P = 0.019) normalized to the TBP reference gene were significantly higher in malignant compared to non-malignant prostate tissues. Employing receiver-operating characteristic (ROC) analyses, PCA3 was the best single tumor marker with the highest area-under-the-curve (AUC = 0.85). A multivariate logit model for the predictability of the tumor was developed, which employed the relative expression levels of EZH2, PCA3, prostein, and TRPM8 and yielded an AUC of 0.90. CONCLUSIONS: The transcript marker PCA3 is a powerful predictor of primary PCa but the inclusion of EZH2, prostein, and TRPM8 adds even more to the diagnostic power. The finding of a significantly higher mRNA expression of three different genes (prostein, PSA, TRPM8) in organ-confined tumors compared to non-organ-confined tumors as well as the multi-marker PCa prediction model developed in the retrospective model system on prostatectomies could be of clinical importance for diagnostic purposes, and should be verified in diagnostic biopsies.
Assuntos
Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Testes Genéticos/métodos , Neoplasias da Próstata/genética , Idoso , Antígenos de Neoplasias/genética , Biomarcadores Tumorais/genética , Biópsia , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/genética , Proteína Potenciadora do Homólogo 2 de Zeste , Humanos , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Análise Multivariada , Complexo Repressor Polycomb 2 , Valor Preditivo dos Testes , Neoplasias da Próstata/patologia , Neoplasias da Próstata/terapia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Canais de Cátion TRPM/genética , Fatores de Transcrição/genética , Falha de TratamentoRESUMO
Chlorinated natural products include vancomycin and cryptophycin A. Their biosynthesis involves regioselective chlorination by flavin-dependent halogenases. We report the structural characterization of tryptophan 7-halogenase (PrnA), which regioselectively chlorinates tryptophan. Tryptophan and flavin adenine dinucleotide (FAD) are separated by a 10 angstrom-long tunnel and bound by distinct enzyme modules. The FAD module is conserved in halogenases and is related to flavin-dependent monooxygenases. On the basis of biochemical studies, crystal structures, and by analogy with monooxygenases, we predict that FADH2 reacts with O2 to make peroxyflavin, which is decomposed by Cl-. The resulting HOCl is guided through the tunnel to tryptophan, where it is activated to participate in electrophilic aromatic substitution.