Molecular Biology

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    Protocols in Current Issue
    Super-resolution Microscopy-based Bimolecular Fluorescence Complementation to Study Protein Complex Assembly and Co-localization
    Authors:  Jingjing Chen, Zulin Yu, Jay R. Unruh, Brian D. Slaughter and Sue L. Jaspersen, date: 02/20/2020, view: 539, Q&A: 0
    [Abstract] Numerous experimental approaches exist to study interactions between two subunits of a large macromolecular complex. However, most methods do not provide spatial and temporal information about binding, which are critical for dissecting the mechanism of assembly of nanosized complexes in vivo. While recent advances in super-resolution ...
    Assessing Self-interaction of Mammalian Nuclear Proteins by Co-immunoprecipitation
    Authors:  Claudia Cattoglio, Iryna Pustova, Xavier Darzacq, Robert Tjian and Anders S. Hansen, date: 02/20/2020, view: 619, Q&A: 0
    [Abstract] Protein-protein interactions constitute the molecular foundations of virtually all biological processes. Co-immunoprecipitation (CoIP) experiments are probably the most widely used method to probe both heterotypic and homotypic protein-protein interactions. Recent advances in super-resolution microscopy have revealed that several nuclear proteins ...
    Surface Plasmon Resonance Analysis of the Protein-protein Binding Specificity Using Autolab ESPIRIT
    Authors:  Pragyan Parimita Rath, Gaurav Anand and Shalini Agarwal, date: 02/20/2020, view: 295, Q&A: 0
    [Abstract] Direct protein-protein interactions are known to regulate a wide range of cellular activities. To understand these contacts one can employ various experimental methods like Dynamic Light Scattering (DLS), Fluorescence Resonance Energy Transfer (FRET), Isothermal titration calorimetry (ITC), Chemical crosslinking, Co-immunoprecipitation (Co-IP), ...
    Identification of Heteroreceptors Complexes and Signal Transduction Events Using Bioluminescence Resonance Energy Transfer (BRET)
    Authors:  Irene Reyes-Resina, Jasmina Jiménez, Gemma Navarro and Rafael Franco, date: 10/05/2019, view: 619, Q&A: 0
    [Abstract] Detecting protein-protein interactions by co-immunoprecipitation provided a major advancement in the immunology research field. In the G-protein-coupled receptors (GPCRs) research field, colocalization and co-immunoprecipitation were used to detect interactions, but doubts arose due to specificity of the antibodies (monoclonal in the case of ...
    Fluorescence Microscopy Assay to Measure HIV-1 Capsid Uncoating Kinetics in vitro
    [Abstract] The stability of the HIV-1 capsid and the spatiotemporal control of its disassembly, a process called uncoating, need to be finely tuned for infection to proceed. Biochemical methods for measuring capsid lattice disassembly in bulk are unable to resolve intermediates in the uncoating reaction. We have developed a single-particle fluorescence ...
    Dynamic and Sequential Protein Reconstitution on Negatively Curved Membranes by Giant Vesicles Fusion
    Authors:  Nicola de Franceschi, Maryam Alqabandi, Winfried Weissenhorn and Patricia Bassereau, date: 07/05/2019, view: 1024, Q&A: 0
    [Abstract] In vitro investigation of the interaction between proteins and positively curved membranes can be performed using a classic nanotube pulling method. However, characterizing protein interaction with negatively curved membranes still represents a formidable challenge. Here, we describe our recently developed approach based on ...
    Tandem Affinity Purification of SBP-CBP-tagged Type Three Secretion System Effectors
    Authors:  Laura Berneking, Marie Schnapp, Theresa Nauth and Moritz Hentschke, date: 06/20/2019, view: 1140, Q&A: 0
    [Abstract] Identification of protein-protein interactions of bacterial effectors and cellular targets during infection is at the core to understand how bacteria manipulate the infected host to overcome the immune response. Potential interacting proteins might be identified by genetic methods, i.e., two hybrid screens and could be verified by ...
    Biofilm Assays on Fibrinogen-coated Silicone Catheters and 96-well Polystyrene Plates
    Authors:  Cristina Colomer-Winter, José A. Lemos and Ana L. Flores-Mireles, date: 03/20/2019, view: 1926, Q&A: 0
    [Abstract] Biofilm formation is a well-known bacterial strategy that protects cells from hostile environments. During infection, bacteria found in a biofilm community are less sensitive to antibiotics and to the immune response, often allowing them to colonize and persist in the host niche. Not surprisingly, biofilm formation on medical devices, such as ...
    In situ, Cell-free Protein Expression on Microarrays and Their Use for the Detection of Immune Responses
    [Abstract] Until recently, whole-proteome microarrays for comprehensive studies of protein interactions were mostly produced by individual cloning and cellular expression of very many open reading frames, followed by protein isolation and purification as well as array production. To overcome this cumbersome process, we have developed a method to generate ...
    Cluster Analysis of Endogenous HER2 and HER3 Receptors in SKBR3 Cells
    [Abstract] The Human Epidermal Growth Factor Receptor (HER) family of receptor tyrosine kinases consists of four, single pass, transmembrane receptor homologs (HER1-4) that act to regulate many critical processes in normal and tumor cells. HER2 is overexpressed in many tumors, and the deregulated proliferation of cancerous cells is driven by cooperation with ...



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