Description
Book SynopsisThis volume describes methods and protocols for the fragment-based screening of proteins using Surface Plasmon Resonance (SPR). The initial chapter of the book discusses the principle of SPR for the identification of biomolecular interactions, while the subsequent chapters introduce methods for labelling proteins with different tags including, histidine and biotin tags. It also discusses techniques and factors that affect the amine and biotin-streptavidin coupling and methods to optimize the interactions. Next, it describes fragment preparation for screening in SPR and presents methods to calculate equilibrium dissociation constant (KD) and ligand efficiency (LE). It reviews techniques of next-generation injections that improve the efficiency of the characterization process over traditional SPR by determining the kinetics and affinity in a single step. Finally, the book elucidates a comprehensive yet representative description of challenges associated with the molecular interaction of proteins using SPR.
Table of ContentsChapter 1_Introduction to SPR.-
Chapter 2_ Principle of SPR (OneStep).-
Chapter 3_Identification of Fragment library.-
Chapter 4_Preparation of protein with Histidine Tag.-
Chapter 5_Preparation of protein with Biotin Tag.-
Chapter 6_Protein stability using Thermal Shift Assay (TSA) - pH.-
Chapter 7_Protein stability using Thermal Shift Assay (TSA) – DMSO.-
Chapter 8_Preparation of Sensor.-
Chapter 9_ Optimization and validation of Amine coupling.-
Chapter 10_ Optimization and validation of Biotin-strepavidin coupling.-
Chapter 11_ Preparation of Fragment for Screening in SPR.-
Chapter 12_ Screening of Fragment in SPR.-
Chapter 13_ Analysis of Data.-
Chapter 14_ Identification and arraying of HITS.-
Chapter 15_ Confirmation of Hits – Kinetics (kon, koff, KD, LE).
