RESUMO
Thiol-containing nucleophiles such as cysteine react spontaneously with the citric acid cycle intermediate fumarate to form S-(2-succino)-adducts. In Bacillus subtilis, a salvaging pathway encoded by the yxe operon has recently been identified for the detoxification and exploitation of these compounds as sulfur sources. This route involves acetylation of S-(2-succino)cysteine to N-acetyl-2-succinocysteine, which is presumably converted to oxaloacetate and N-acetylcysteine, before a final deacetylation step affords cysteine. The critical oxidative cleavage of the C-S bond of N-acetyl-S-(2-succino)cysteine was proposed to depend on the predicted flavoprotein monooxygenase YxeK. Here, we characterize YxeK and verify its role in S-(2-succino)-adduct detoxification and sulfur metabolism. Detailed biochemical and mechanistic investigation of YxeK including 18 O-isotope-labeling experiments, homology modeling, substrate specificity tests, site-directed mutagenesis, and (pre-)steady-state kinetics provides insight into the enzyme's mechanism of action, which may involve a noncanonical flavin-N5-peroxide species for C-S bond oxygenolysis.
Assuntos
Cisteína/análogos & derivados , Cisteína/genética , Flavoproteínas/genética , Oxigenases de Função Mista/genética , Acetilação , Bacillus subtilis/genética , Bacillus subtilis/metabolismo , Cisteína/metabolismo , Flavinas/genética , Flavinas/metabolismo , Flavoproteínas/metabolismo , Fumaratos/metabolismo , Cinética , Modelos Químicos , Mutagênese Sítio-Dirigida , Óperon/genética , Especificidade por Substrato/genética , Compostos de Sulfidrila/metabolismoRESUMO
The expression of neuroendocrine marker proteins in undifferentiated colorectal cancers has not yet been studied in great detail. Therefore, the survival of 20 patients with small cell undifferentiated colorectal cancers treated at our institution between 1982 and 1997 (0.8% of all operated colorectal carcinomas) was correlated with the extent of neuroendocrine differentiation. Chromogranin A, synaptophysin, syntaxin1, VAMP2, SNAP25, and alpha/beta-SNAP were used as neuroendocrine markers. Based on the degree of immunoreactivity for these marker proteins, tumors were divided into group 0 (<2% cells stained positive for neuroendocrine markers) and group 1 (>2% cells stained positive). Patients were followed up for at least 5 years or until death. Nine of twenty (45%) undifferentiated colorectal tumors expressed neuroendocrine markers (group 1). Only one patient of this group survived 2 years (11%), whereas the 2-year survival rate was 45.4% in group 0. Nine of eleven patients of group 0 were diagnosed in UICC stage I-III, whereas eight of nine tumors with expression of neuroendocrine markers were diagnosed in UICC stage IV (P = 0.002). Our results show that neuroendocrine differentiation is often seen in small cell undifferentiated colorectal cancer. It correlates with a more aggressive course of the disease.
Assuntos
Biomarcadores Tumorais , Neoplasias Colorretais/patologia , Tumores Neuroendócrinos/patologia , Proteínas de Transporte Vesicular , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Superfície/metabolismo , Proteínas de Transporte/metabolismo , Diferenciação Celular , Neoplasias Colorretais/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Masculino , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/metabolismo , Tumores Neuroendócrinos/metabolismo , Prognóstico , Proteínas R-SNARE , Proteínas SNARE , Proteínas de Ligação a Fator Solúvel Sensível a N-Etilmaleimida , Sinaptofisina/metabolismo , Proteína 25 Associada a Sinaptossoma , Sintaxina 1RESUMO
The frequency and prognostic significance of neuroendocrine marker expression in undifferentiated colorectal cancers has not yet been studied in great detail. Therefore, the survival of 20 patients with small cell undifferentiated colorectal cancers, treated at our institution between 1982 and 1997 (0.8% of all operated colorectal carcinomas), was correlated with the extent of neuroendocrine differentiation. Chromogranin A, synaptophysin, syntaxin1, VAMP2, SNAP25 and alpha/beta-SNAP were used as neuroendocrine markers. Based on the degree of immunoreactivity for these marker proteins, tumors were separated into group 0 (<2% cells stained positive for neuroendocrine markers) and group 1 (>2% cells stained positive). Patients were followed up for at least 5 years or until death. Of 20 (45%) undifferentiated colorectal tumors, 9 expressed neuroendocrine markers (group 1). Only one patient of this group survived for 2 years (11%), whereas the 2-year-survival rate was 45.4% in group 0. Of the 11 patients in group 0, 9 were diagnosed with UICC stages I-III, whereas 8 of 9 tumors with expression of neuroendocrine markers were diagnosed with UICC stage IV ( P=0.002). Our results show that neuroendocrine differentiation is often seen in small cell undifferentiated colorectal cancer. It correlates with a more aggressive course of the disease.
