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Trizma® base

(NH 4)2 SO4

Company: Sigma-Aldrich
Catalog#: T6066
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Two Different Methods of Quantification of Oxidized Nicotinamide Adenine Dinucleotide (NAD+) and Reduced Nicotinamide Adenine Dinucleotide (NADH) Intracellular Levels: Enzymatic Coupled Cycling Assay and Ultra-performance Liquid Chromatography (UPLC)-Mass Spectrometry
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Date:
2018-07-20
[Abstract]  Current studies on the age-related development of metabolic dysfunction and frailty are each day in more evidence. It is known, as aging progresses, nicotinamide adenine dinucleotide (NAD+) levels decrease in an expected physiological process. Recent studies have shown that a reduction in NAD+ is a key factor for the development of age-associated metabolic decline. Increased NAD+ levels in vivo results in activation of pro-longevity and health span-related factors. Also, it improves several physiological and metabolic parameters of aging, including muscle function, exercise capacity, glucose tolerance, and cardiac function in mouse models of natural and accelerated aging.

Given the importance of monitoring cellular NAD+ and ...
[摘要]  目前关于代谢功能障碍和虚弱的年龄相关发展的研究每天都有更多的证据。众所周知,随着衰老的进展,烟酰胺腺嘌呤二核苷酸(NAD + )水平在预期的生理过程中降低。最近的研究表明,NAD + 的减少是与年龄相关的代谢衰退发展的关键因素。增加NAD + 水平体内导致激活寿命和健康跨度相关因素。此外,它改善了老化的几个生理和代谢参数,包括自然和加速老化的小鼠模型中的肌肉功能,运动能力,葡萄糖耐量和心脏功能。

鉴于监测细胞NAD + 和NADH水平的重要性,有一个值得信赖的方法是至关重要的。该方案的目的是在有效且广泛适用的测定中描述来自组织和细胞的NAD + 和NADH提取以及其图形和定量分析。

【背景】氧化烟酰胺腺嘌呤二核苷酸和还原型烟酰胺腺嘌呤二核苷酸(NAD + 和NADH)是重要的生物辅助因子,它们在几种合成代谢和分解代谢功能中提供和接受电子。它们参与诸如糖酵解,三羧酸循环和氧化磷酸化的反应。此外,它还作为DNA损伤修复中涉及的几种酶的底物,例如sirtuins和poly(ADP-核糖)聚合酶(PARP)(Imai和Guarente,2014; Verdin,2015; Yoshino et al。 ,2018)。

NAD + ...

Measuring CD38 Hydrolase and Cyclase Activities: 1,N6-Ethenonicotinamide Adenine Dinucleotide (ε-NAD) and Nicotinamide Guanine Dinucleotide (NGD) Fluorescence-based Methods
Author:
Date:
2018-07-20
[Abstract]  CD38 is a multifunctional enzyme involved in calcium signaling and Nicotinamide Adenine Dinucleotide (NAD+) metabolism. Through its major activity, the hydrolysis of NAD+, CD38 helps maintain the appropriate levels of this molecule for all NAD+-dependent metabolic processes to occur. Due to current advances and studies relating NAD+ decline and the development of multiple age-related conditions and diseases, CD38 gained importance in both basic science and clinical settings. The discovery and development of strategies to modulate its function and, possibly, treat diseases and improve health span put CD38 under the spotlights. Therefore, a consistent and reliable method to measure its activity and explore its use in medicine is required. We ... [摘要]  CD38是参与钙信号传导和烟酰胺腺嘌呤二核苷酸(NAD + )代谢的多功能酶。通过其主要活性,NAD + 的水解,CD38有助于维持该分子的适当水平,从而发生所有NAD + 依赖性代谢过程。由于目前有关NAD + 下降和多种年龄相关疾病和疾病发展的研究和进展,CD38在基础科学和临床环境中都具有重要意义。调整其功能,可能治疗疾病和改善健康状况的战略的发现和发展使CD38成为焦点。因此,需要一种一致且可靠的方法来测量其活性并探索其在医学中的用途。我们在这里描述了我们的小组如何测量CD38的水解酶和环化酶活性的方法,利用基于荧光的酶测定法在平板阅读器中使用1,N 6 -Ethenonicotinamide Adenine Dinucleotide(ε-NAD)进行)和烟酰胺鸟嘌呤二核苷酸(NGD)分别作为底物。

【背景】目前关于代谢功能障碍和虚弱的年龄相关发展的研究每天都有更多的证据。众所周知,随着老化的进行,NAD + 水平在预期过程中降低。最近的研究表明,烟酰胺腺嘌呤二核苷酸(NAD + )的减少与年龄相关的代谢下降有关(Massudi et al。,2012)。增加NAD + 水平体内,导致延长寿命和健康跨度相关因素的活化,并改善衰老的几个生理和代谢参数(Camacho-Pereira ...

Guanine Nucleotide Exchange Assay Using Fluorescent MANT-GDP
Author:
Date:
2018-04-05
[Abstract]  GTPases are molecular switches that cycle between the inactive GDP-bound state and the active GTP-bound state. GTPases exchange nucleotides either by its intrinsic nucleotide exchange or by interaction with guanine nucleotide exchange factors (GEFs). Monitoring the nucleotide exchange in vitro, together with reconstitution of direct interactions with regulatory proteins, provides key insights into how a GTPase is activated. In this protocol, we describe core methods to monitor nucleotide exchange using fluorescent N-Methylanthraniloyl (MANT)-guanine nucleotide. [摘要]  GTP酶是分子开关,在无效GDP结合状态和活性GTP结合状态之间循环。 GTP酶通过其内在的核苷酸交换或通过与鸟嘌呤核苷酸交换因子(GEF)的相互作用来交换核苷酸。 监测体外核苷酸交换,以及与调节蛋白直接相互作用的重构,为GTP酶如何被激活提供了重要见解。 在该协议中,我们描述了使用荧光N-甲基呋喃酰基(MANT) - 鸟嘌呤核苷酸来监测核苷酸交换的核心方法。

【背景】GTPase是鸟嘌呤核苷酸结合蛋白,调节细胞过程的广度,从蛋白质生物合成到细胞周期进展,从细胞骨架重组到膜运输。 GTPases可以被认为是分子开关,它在GDP结合“关闭”状态和GTP结合“开启”状态之间循环;在通过GTP的GDP核苷酸交换结合GTP时,GTP酶变得活跃并且将结合下游效应蛋白以招募和激活这些效应子的生物学功能。 GTP酶通过与开关I环(G2结构域)的高度保守苏氨酸和开关II环(G3结构域)的DxxG基序内的甘氨酸的相互作用结合GTP的γ-磷酸。 GTP水解后,与γ-磷酸相互作用的丧失导致动态构象变化,从而使GTPase变为关闭状态(Vetter and ...

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