Quantification of ERK Kinase Activity in Biological Samples Using Differential Sensing.
Quantification of ERK Kinase Activity in Biological Samples Using Differential Sensing.
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使用差分传感定量生物样品中的 ERK 激酶活性。
DOI：
10.1021/acschembio.9b00580
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发表时间：
2020
影响因子：
4
通讯作者：
Dalby,KevinN
中科院分区：
文献类型：
--
作者：
Zamora-Olivares,Diana;Kaoud,TamerS;Zeng,Lingyu;Pridgen,JaceyR;Zhuang,DeborahL;Ekpo,YakndaraE;Nye,JessicaR;Telles,Mitchell;Anslyn,EricV;Dalby,KevinN
理解复杂的生物系统需要有能力评估基因、蛋白质和细胞反应的相互作用网络。支持快速量化这些网络的赋能技术将促进生物模型的开发，并有助于确定治疗目标和评估治疗计划。蛋白质磷酸化的生化过程是细胞信号传导的几乎所有方面的基础，通常通过免疫印迹方法（Western印迹）或最近的蛋白质组学方法进行评估，其提供细胞中蛋白质浓度及其修饰的定性估计。然而，蛋白质修饰很难与活性相关，虽然免疫印迹和蛋白质组学方法具有定量的潜力，但它们需要一系列复杂的步骤，从而降低了重现性。在这里，提出了一种互补的方法，允许快速定量细胞裂解物和肿瘤样品中的蛋白激酶活性。使用细胞ERK（细胞外信号调节激酶）的活性作为测试案例，使用阵列传感方法，利用基于差异肽的生物传感器和化学计量工具的库，用于快速定量微克未分级细胞裂解物和肿瘤提取物中的纳克活性ERK。这种方法具有高通量和定量单一生物样品中多种蛋白激酶活性的潜力。与其他方法相比，这种差异传感方法的关键优势在于，它消除了添加外源抑制剂以抑制主要脱靶激酶活性的需要，并允许我们定量测试样品中活性激酶的量，而不是测量诱导或抑制后其活性的变化。
The understanding of complex biological systems requires an ability to evaluate interacting networks of genes, proteins, and cellular reactions. Enabling technologies that support the rapid quantification of these networks will facilitate the development of biological models and help to identify treatment targets and to assess treatment plans. The biochemical process of protein phosphorylation, which underlies almost all aspects of cell signaling, is typically evaluated by immunoblotting procedures (Western blot) or more recently proteomics procedures, which provide qualitative estimates of the concentration of proteins and their modifications in cells. However, protein modifications are difficult to correlate with activity, and while immunoblotting and proteomics approaches have the potential to be quantitative, they require a complex series of steps that diminish reproducibility. Here, a complementary approach is presented that allows for the rapid quantification of a protein kinase activity in cell lysates and tumor samples. Using the activity of cellular ERK (extracellular signal-regulated kinase) as a test case, an array sensing approach that utilizes a library of differential peptide-based biosensors and chemometric tools was used to rapidly quantify nanograms of active ERK in micrograms of unfractionated cell lysates and tumor extracts. This approach has the potential both for high-throughput and for quantifying the activities of multiple protein kinases in a single biological sample. The critical advantages of this differential sensing approach over others are that it removes the need for the addition of exogenous inhibitors to suppress the activities of major off-target kinases and allows us to quantitate the amount of active kinase in tested samples rather than measuring the changes in its activity upon induction or inhibition.
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对类风湿关节炎患者基于计算机的教育课程的评估。
DOI：
--
发表时间：
1985
期刊：
The Journal of rheumatology
影响因子：
--
作者：
Wetstone,SL;Sheehan,TJ;Votaw,RG;Peterson,MG;Rothfield,N
通讯作者：
Rothfield,N
影响因子：
12.7
作者：
MORISKY, DE;LEVINE, DM;SMITH, CR
通讯作者：
SMITH, CR
影响因子：
5.4
作者：
L. Bradley;L. D. Young;K. Anderson;L. McDaniel;R. Turner;C. Agudelo
通讯作者：
C. Agudelo
DOI：
--
发表时间：
1989
期刊：
影响因子：
--
作者：
M. Stewart;D. Roter
通讯作者：
D. Roter
改善患者对关节炎治疗方案的配合。
影响因子：
--
作者：
Alan M. Jette
通讯作者：
Alan M. Jette