化学药中微生物相关及生化特性检测方案

智能文字提取功能测试中

Biolin Scientific[Progress Together] [Application Overview]3 Improving drug discovery using QCM-D Screening of compound interactions with cells and protein drug targets is often essential in drug discovery. Quartz Crystal Microbalance with Dissipation monitoring， QCM-D， enables such measurements in real-time. Introduction In particular， the ability of the QCM-D technique to senseconformational and structural changes in molecular layers orwhole cells on the sensor surface using its unique ability toprobe changes in softness (energy dissipation) provides a majoradvantage. These capabilities have potential within drug discovery，enabling both detection of potential new pharmaceuticalcandidates in drug target screening assays， and improvedunderstanding of target interactions mechanisms. [Figure 1]： Illustration of carcinoma cells on a QCM-D sensor， and the exposureto EGF.Addition of EGF causes changes in the cytoskeleton. Cellular responses to growth factors in humancarcinoma cells [1] QCM-D provides a sensitive platform for cell-surface interactionswhich can be monitored in real-time. Here， human carcinomacells attached to the sensor ex-situ and their interaction withthe epidermal growth factor (EGF) was analyzed [1] (Fig.1). Thebinding of EGF to the EGF receptor (EGFR) results in cytoskeletonrearrangements in the cells which are successfully probed byQCM-D as changes in the viscoelasticity of the cell layer. [Figure 2]： Plasminogen protein conformational change due to small moleculeinteraction. Protein conformational changes caused by smallcompounds [2][3] Several publications have highlighted QCM-D as a useful toolin protein drug target screening. These assays depend onconformational changes to the tertiary structure of the proteinsupon binding of small compounds. Here， the unfolding ofthe protein plasminogen was probed upon addition of lysineanalogues [2] (Fig.2). Similarly，conformational re-arrangementshave been studied in the HIV-1 envelope protein [3]. The His-tagged recombinant HIV-1 envelope protein was immobilized onthe QCM-D sensor and its interactions with different compoundswere followed in real-time. In both studies changes in the energydissipation proved crucial for understanding conformationalevents. [Figure 3]： Outline of the estrogen-DNA binding assay and structure of the twoligands. Understanding biomolecular interactions and processes [4][5] Insights into protein-protein or protein-DNA interactions are ofvital importance in drug development.Conformational effects innuclear receptors， such as the estrogen receptor， is one relevantexample [4]. Upon ligand binding the receptors bind to DNA.By immobilization of biotinylated DNA， followed by injectionof estrogen receptors in the absence or presence of ligands，the interactions can be studied by QCM-D [4]. Also， processesof self-oligomerization and aggregation of proteins involved in human protein misfolding diseases can be studied using QCM-D.Measurements revealed that oligomers formed by the Alzheimer'sdisease (AD) associated peptide AB are "softer" than themonomers upon interacting with a model surface [5]， somethingthat might be related to their toxicity in AD. ( References： ) ( [1] Chen， J.Y.， Minghong L i ， P enn， L.S.， and X i， J. Real-Time and L abel-Free D etecti o n of Cellular Response to Signaling Mediate d by D i stinct Subclasses of Epidermal Growth Factor Receptors， Anal. Chem.， 83， 3 1 41 - 3146 (2011). ) ( [2] Nileback，E . ， Westberg，F.， D einum， J . ， a nd Svedhem， S. V i scoelastic Sensing of Conformational Changes in Plasminogen Induced upon Binding of LowMolecular Weight Compounds， Anal. Chem.， 82(20)， 83 7 4-6(2010). ) ( [3] Lee， H-S & Contarino， M . ， M . Um a shankara， Schon ， A.， Freir e ， E.， Smith Ill，A.B.， Chaiken，I . M .， and P enn， L . S. Use of the quartz crystal m icrobalance t o m onitor ligand-induced conformational rearrangements in HIV-1envelope p rotein gp 120， Anal. Bioanal. Chem.， 396， 1 1 43-1152(2010). ) ( [4] Wendy YX Peh， W.X.Y.， R e imhult， E. ， Teh， H. F .， T homsen，J.S.， a n d Su， X. Understandin g ligand binding effects on the conformation of estrogen recep- tor alpha-DNA comp le xes： A c o mbinational qua rt z c ryst al microbalance withdissipatio n and surface plasmon resonance study， Biophys. J .， 92， 4415-4423 (2007). ) ( [5] Pereira MC， S.A.M.， and B rezesinski G . ， I s the viscoela s ticity o f Alz- h eimer's A beta42 pe p ti de o l ig o mers a g en eral p r o perty o f protein oligomers r elated to their toxicity ， Langmuir， 26( 1 4)， 12060-12067(201 0 ). ) []Q-Sense AO Biolin Scientific[]Q-Sense AO