Current protocols in chemical biology最新文献_第4页

Enzymatic Synthesis of Backbone-Modified Oligonucleotides Using T4 DNA Ligase 利用T4 DNA连接酶合成骨架修饰寡核苷酸

Current protocols in chemical biology Pub Date : 2019-01-28 DOI: 10.1002/cpch.62

Donaat Kestemont, Piet Herdewijn, Marleen Renders

引用次数: 3

Native Chemical Ligation of Peptides and Proteins 多肽和蛋白质的天然化学连接

Current protocols in chemical biology Pub Date : 2019-01-15 DOI: 10.1002/cpch.61

Philip A. Cistrone, Michael J. Bird, Dillon T. Flood, Anthony P. Silvestri, Jordi C. J. Hintzen, Darren A. Thompson, Philip E. Dawson

引用次数: 16

In Vivo Identification of Protein Kinase Substrates by Kinase-Oriented Substrate Screening (KIOSS) 激酶导向底物筛选(KIOSS)在体内鉴定蛋白激酶底物

Current protocols in chemical biology Pub Date : 2019-01-07 DOI: 10.1002/cpch.60

Tomoki Nishioka, Mutsuki Amano, Yasuhiro Funahashi, Daisuke Tsuboi, Yukie Yamahashi, Kozo Kaibuchi

{"title":"In Vivo Identification of Protein Kinase Substrates by Kinase-Oriented Substrate Screening (KIOSS)","authors":"Tomoki Nishioka, Mutsuki Amano, Yasuhiro Funahashi, Daisuke Tsuboi, Yukie Yamahashi, Kozo Kaibuchi","doi":"10.1002/cpch.60","DOIUrl":"10.1002/cpch.60","url":null,"abstract":"Protein phosphorylation plays a critical role in the regulation of cellular function. Information on protein phosphorylation and the responsible kinases is important for understanding intracellular signaling. A method for in vivo screening of kinase substrates named KIOSS (kinase-oriented substrate screening) has been developed. This protocol provides a method that utilizes phosphoprotein-binding modules such as 14-3-3 protein, the pin1-WW domain, and the chek2-FHA domain as biological filters to successfully enrich phosphorylated proteins related to intracellular signaling rather than housekeeping and/or structural proteins. More than 1000 substrate candidates for PKA, PKC, MAPK, and Rho-kinase in HeLa cells, as well as phosphorylation downstream of D1R, NMDAR, adenosine A2a receptor, PKA, PKC, MAPK, and Rho-kinase in mouse brain slice cultures have been identified by this method. An online database named KANPHOS (Kinase-Associated Neural Phospho-Signaling) provides the phosphorylation signals identified by these studies, as well as those previously reported in the literature. © 2019 by John Wiley & Sons, Inc.","PeriodicalId":38051,"journal":{"name":"Current protocols in chemical biology","volume":"11 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2019-01-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpch.60","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"36828869","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 10

Engineering Bacterial Shape Using Soft Matter Microchambers 利用软物质微室设计细菌形状

Current protocols in chemical biology Pub Date : 2018-12-07 DOI: 10.1002/cpch.59

Lars David Renner

引用次数: 2

Quantification of Cellular Proteostasis in Live Cells by Fluorogenic Assay Using the AgHalo Sensor 用AgHalo传感器荧光测定活细胞中细胞蛋白酶的含量

Current protocols in chemical biology Pub Date : 2018-11-29 DOI: 10.1002/cpch.58

Matthew Fares, Xin Zhang

{"title":"Quantification of Cellular Proteostasis in Live Cells by Fluorogenic Assay Using the AgHalo Sensor","authors":"Matthew Fares, Xin Zhang","doi":"10.1002/cpch.58","DOIUrl":"10.1002/cpch.58","url":null,"abstract":"Proper cellular proteostasis is essential to cellular fitness and viability. Exogenous stress conditions compromise proteostasis and cause aggregation of cellular proteins. We have developed a fluorogenic sensor (AgHalo) to quantify stress-induced proteostasis deficiency. The AgHalo sensor uses a destabilized HaloTag variant to represent aggregation-prone cellular proteins and is equipped with a series of fluorogenic probes that exhibit a fluorescence increase when the sensor forms either soluble oligomers or insoluble aggregates. Herein, we present protocols that describe how the AgHalo sensor can be employed to visualize and quantify proteome stress in live cells using a direct fluorescence read-out and visualization with a fluorescence microplate reader and a microscope. Additionally, protocols for using the AgHalo sensor in combination with fluorogenic probes and commercially available HaloTag probes to enable two-color imaging experiments are described. These protocols will enable use of the AgHalo sensor to visualize and quantify proteostasis in live cells, a task that is difficult to accomplish using previous, always-fluorescent methods. © 2018 by John Wiley & Sons, Inc.","PeriodicalId":38051,"journal":{"name":"Current protocols in chemical biology","volume":"11 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-11-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpch.58","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"36732552","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Issue Information TOC 发布信息TOC

Current protocols in chemical biology Pub Date : 2018-11-26 DOI: 10.1002/cpch.53

引用次数: 0

Endotoxin-Free Preparation of Graphene Oxide and Graphene-Based Materials for Biological Applications 无内毒素氧化石墨烯制备及生物应用石墨烯基材料

Current protocols in chemical biology Pub Date : 2018-10-04 DOI: 10.1002/cpch.51

Dorsa Parviz, Michael Strano

{"title":"Endotoxin-Free Preparation of Graphene Oxide and Graphene-Based Materials for Biological Applications","authors":"Dorsa Parviz, Michael Strano","doi":"10.1002/cpch.51","DOIUrl":"10.1002/cpch.51","url":null,"abstract":"Due to their two-dimensional structure and unique properties, graphene and its derivatives have been extensively studied for their potential applications in various fields ranging from electronics to composites. Particularly, their high surface area, electrical conductivity, mechanical strength, dispersability in aqueous phase, and possibility of surface modification make them promising candidates for biomedical applications including biosensing, drug delivery, tissue engineering, cell imaging, and therapeutics. The functioning of graphene nanosheets in these applications is dependent on their structure and properties, which are mainly determined by their preparation and processing methods. Exfoliation techniques are the most common methods for preparation of graphene nanosheets for biomedical applications due to their high yield and scalability. Further modification of these methods is necessary to produce biocompatible and toxin-free graphene that can be safely incorporated into biological media. Here, we describe protocols for chemical and mechanical exfoliation of graphite to produce endotoxin-free and highly stable graphene oxide and graphene dispersions. Additional protocols are provided for proper pre- and post-processing of nanosheets and endotoxin measurement techniques. © 2018 by John Wiley & Sons, Inc.","PeriodicalId":38051,"journal":{"name":"Current protocols in chemical biology","volume":"10 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-10-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpch.51","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"36557056","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 11

A Sensitive High-Throughput Screening Method for Identifying Small Molecule Stimulators of the Core Particle of the Proteasome 一种鉴定蛋白酶体核心颗粒小分子刺激物的灵敏高通量筛选方法

Current protocols in chemical biology Pub Date : 2018-10-04 DOI: 10.1002/cpch.52

Rachel A. Coleman, Darci J. Trader

引用次数: 4

In Vivo Evaluation of Ligand Targeted Drug Conjugates for Cancer Therapy 配体靶向药物偶联物治疗癌症的体内评价

Current protocols in chemical biology Pub Date : 2018-09-13 DOI: 10.1002/cpch.49

Mena Asha Krishnan, Amit Pandit, Venkatesh Chelvam

{"title":"In Vivo Evaluation of Ligand Targeted Drug Conjugates for Cancer Therapy","authors":"Mena Asha Krishnan, Amit Pandit, Venkatesh Chelvam","doi":"10.1002/cpch.49","DOIUrl":"10.1002/cpch.49","url":null,"abstract":"The development of small molecule ligand–targeted therapeutics is currently of paramount importance for treatment of cancer due to their potential to reduce system toxicity and increase potency of a delivered chemotherapeutic drug. The main aim of a targeted drug-delivery technique is to release the drug cargo selectively into tumor tissues, avoiding off-site toxicity to healthy tissues and organs during chemotherapy. In this strategy, a chemotherapeutic drug is conjugated to a homing ligand, which has high affinity for proteins over-expressed on cancer cells, via a peptide linker and a self-immolative segment that facilitates intracellular release of drug cargo. During development of targeted drug conjugates, preclinical evaluation in tumor models of small animals like mice adds valuable data on the clinical performance of the drug. This article contains a set of protocols for implantation of tumor, determination of optimum dosage required for effective treatment, and estimation of maximum tolerated dose required for any visible side effects during treatment of cancer in tumor models of mice. © 2018 by John Wiley & Sons, Inc.","PeriodicalId":38051,"journal":{"name":"Current protocols in chemical biology","volume":"10 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpch.49","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"36489172","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Preparation of Ligand-Targeted Drug Conjugates for Cancer Therapy and Their Evaluation In Vitro 肿瘤治疗配体靶向药物偶联物的制备及其体外评价

Current protocols in chemical biology Pub Date : 2018-09-13 DOI: 10.1002/cpch.50

Mena Asha Krishnan, Sagnik Sengupta, Venkatesh Chelvam

{"title":"Preparation of Ligand-Targeted Drug Conjugates for Cancer Therapy and Their Evaluation In Vitro","authors":"Mena Asha Krishnan, Sagnik Sengupta, Venkatesh Chelvam","doi":"10.1002/cpch.50","DOIUrl":"10.1002/cpch.50","url":null,"abstract":"Present treatment strategies focus on minimizing unwanted toxicity to healthy cells during chemotherapeutic treatment. This is achieved by developing strategies to selectively deliver drugs to malignant cells over-expressing specific protein biomarkers. The drugs are attached via a self-immolative linker to a small molecule homing ligand having affinity for protein biomarkers over-expressed during disease states. Several such targeting-ligand drug conjugates have now reached preclinical and clinical trials, and this article aims to show a general methodology to prepare the same. Using solid-phase peptide synthesis (SPPS) methodology, the targeting ligand is covalently linked to a peptide spacer having appropriate hydrophobic and hydrophilic amino acids. The targeting ligand–attached peptide spacer is next conjugated with the required drug molecule through a cleavable disulfide bond in a solution-phase reaction. This protocol further elucidates the step-by-step procedures to be followed for complete evaluation of newly synthesized ligand-targeted drug conjugates in vitro. © 2018 by John Wiley & Sons, Inc.","PeriodicalId":38051,"journal":{"name":"Current protocols in chemical biology","volume":"10 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2018-09-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/cpch.50","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"36489827","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1