RESUMEN
Significant attention has been shifted toward the use and development of biodegradable polymeric materials to mitigate environmental accumulation and potential health impacts. One such material, poly(aspartic acid) (PAA), is a biodegradable alternative to superabsorbent poly(carboxylates), like poly(acrylate). Three enzymes are known to hydrolyze PAA: PahZ1KT-1 and PahZ2KT-1 from Sphingomonas sp. KT-1 and PahZ1KP-2 from Pedobacter sp. KP-2. We previously reported the X-ray crystal structure for PahZ1KT-1, which revealed a homodimer complex with a strongly cationic surface spanning one side of each monomer. Here, we report the first characterization of any polymer hydrolase binding to DNA, where modeling data predict binding of the polyanionic DNA near the cationic substrate binding surface. Our data reveal that PahZ1 homologues from Sphingomonas sp. KT-1 and Pedobacter sp. KP-2 bind ssDNA and dsDNA with nanomolar binding affinities. PahZ1KT-1 binds ssDNA and dsDNA with an apparent dissociation constant, KD,app = 81 ± 14 and 19 ± 1 nM, respectively, and these estimates are similar to the same behaviors exhibited by PahZ1KP-2. Gel permeation chromatography data reveal that dsDNA binding promotes inhibition of PahZ1-catalyzed PAA biodegradation for each homologue. We propose a working model wherein binding of PahZ1 to extracellular biofilm DNA aids in the localization of the hydrolase to the environment in which PAA would first be encountered, thereby providing a mechanism to degrade extracellular PAA and potentially harvest aspartic acid for nutritional uptake.
Asunto(s)
Sphingomonas , Sphingomonas/enzimología , Pedobacter/enzimología , ADN/metabolismo , Hidrolasas/metabolismo , Hidrolasas/química , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Péptidos/metabolismo , Péptidos/química , ADN de Cadena Simple/metabolismo , Modelos Moleculares , Unión Proteica , Ácido Aspártico/metabolismo , Ácido Aspártico/químicaRESUMEN
Autophagy, an autophagosome and lysosome-based eukaryotic cellular degradation system, has previously been implicated in lifespan regulation in different animal models. In this report, we show that expression of the RNAi transgenes targeting the transcripts of the key autophagy genes Atg1 or Atg18 in adult fly muscle or glia does not affect the overall levels of autophagosomes in those tissues and does not change the lifespan of the tested flies but the lifespan reduction phenotype has become apparent when Atg1 RNAi or Atg18 RNAi is expressed ubiquitously in adult flies or after autophagy is eradicated through the knockdown of Atg1 or Atg18 in adult fly adipocytes. Lifespan reduction was also observed when Atg1 or Atg18 was knocked down in adult fly enteroblasts and midgut stem cells. Overexpression of wild-type Atg1 in adult fly muscle or adipocytes reduces the lifespan and causes accumulation of high levels of ubiquitinated protein aggregates in muscles. Our research data have highlighted the important functions of the key autophagy genes in adult fly adipocytes, enteroblasts, and midgut stem cells and their undetermined roles in adult fly muscle and glia for lifespan regulation.
Asunto(s)
Homólogo de la Proteína 1 Relacionada con la Autofagia , Autofagia , Proteínas de Drosophila , Drosophila melanogaster , Longevidad , Animales , Autofagia/genética , Homólogo de la Proteína 1 Relacionada con la Autofagia/genética , Homólogo de la Proteína 1 Relacionada con la Autofagia/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Longevidad/genética , Interferencia de ARNRESUMEN
OBJECTIVE: To evaluate toxicity and cost-effectiveness of intensity modulated radiation therapy (IMRT) versus 3-dimensional conformal radiation therapy (3DCRT) in the postoperative treatment of uterine and cervical cancer. METHODS: Between 2000 and 2012, eighty patients at our institution received post-hysterectomy 3DCRT (46) or IMRT (34) for uterine or cervical cancer. Baseline characteristics, outcome, and ≥CTCAE grade 2 toxicities were compared between the two groups. Predictors of toxicity-free survival were identified. A decision analysis model was designed to capture individual health states at 1, 2, and 3 years after treatment. Micro-costing technique and estimated quality-adjusted life years (QALYs) were used to calculate incremental cost-effectiveness ratio (ICER). RESULTS: Utilization of IMRT increased from 25% (2005-2007) to 75% (2008-2012). Recurrence-free and overall survival rates were not different between the two groups. Toxicity rates were reduced with IMRT versus 3DCRT (HR 0.42, p=0.04). Women who received IMRT had numerically lower rates of late gastrointestinal and genitourinary toxicity and significantly lower rates of late overall toxicity at 3 years (16% vs. 45%, p=0.04). On univariate analysis, IMRT was associated with decreased late toxicity (HR 0.43, p=0.04). Treatment costs were higher and toxicity costs were lower with IMRT. IMRT had an ICER of $235,233 (year 1), $114,270 (year 2), and $75,555 (year 3) per QALY gained. CONCLUSION: IMRT is associated with reduced late overall toxicity compared to 3DCRT without compromising clinical outcome. IMRT is not cost-effective during the early chronic toxicity phase, but it becomes more cost-effective over time.
Asunto(s)
Análisis Costo-Beneficio , Histerectomía , Radioterapia Conformacional/métodos , Neoplasias Uterinas/radioterapia , Adulto , Anciano , Anciano de 80 o más Años , Bases de Datos Factuales , Técnicas de Apoyo para la Decisión , Femenino , Costos de Hospital/estadística & datos numéricos , Humanos , Persona de Mediana Edad , Periodo Posoperatorio , Años de Vida Ajustados por Calidad de Vida , Radioterapia Adyuvante , Radioterapia Conformacional/efectos adversos , Radioterapia Conformacional/economía , Radioterapia de Intensidad Modulada/efectos adversos , Radioterapia de Intensidad Modulada/economía , Radioterapia de Intensidad Modulada/métodos , Sistema de Registros , Estudios Retrospectivos , Análisis de Supervivencia , Resultado del Tratamiento , Utah , Neoplasias del Cuello Uterino/economía , Neoplasias del Cuello Uterino/mortalidad , Neoplasias del Cuello Uterino/radioterapia , Neoplasias del Cuello Uterino/cirugía , Neoplasias Uterinas/economía , Neoplasias Uterinas/mortalidad , Neoplasias Uterinas/cirugíaRESUMEN
Olanzapine, an atypical antipsychotic drug, was previously shown to protect neuronal cells against nutrient deprivation and to enhance neurite outgrowth. In an effort to identify small molecules with greater potency, the structure of olanzapine was used as a template to search commercially available chemical inventories for compounds with similar features. These compounds were evaluated for their ability to protect cells against glutamine deprivation and low-serum conditions. Positive compounds, 'hits' from initial screening, were then tested for stimulation of neurite outgrowth, alone and in combination with suboptimum concentrations of nerve growth factor (NGF). Numerous neuroprotective compounds (mw < 550 Da) were identified that significantly stimulated neurite outgrowth in PC12 cells. These included 4', 6'-diamidino-2-phenylindole, a nuclear stain; staurosporine, an antibiotic and kinase inhibitor; and 2-phenylamino-adenosine, an adenosine analog. The small molecules were comparable with NGF, and in fact, replaced NGF in outgrowth assays. Pharmacophore analysis of the hits led to the design and synthesis of an active compound, LSU-D84, which represented an initial lead for drug discovery efforts.