Binding Revisited-Avidity in Cellular Function and Signaling.

When characterizing biomolecular interactions, avidity, is an umbrella term used to describe the accumulated strength of multiple specific and unspecific interactions between two or more interaction partners. In contrast to the affinity, which is often sufficient to describe monovalent interactions in solution and where the binding strength can be accurately determined by considering only the relationship between the microscopic association and dissociation rates, the avidity is a phenomenological macroscopic parameter linked to several microscopic events. Avidity also covers potential effects of reduced dimensionality and/or hindered diffusion observed at or near surfaces e.g., at the cell membrane. Avidity is often used to describe the discrepancy or the "extra on top" when cellular interactions display binding that are several orders of magnitude stronger than those estimated in vitro. Here we review the principles and theoretical frameworks governing avidity in biological systems and the methods for predicting and simulating avidity. While the avidity and effects thereof are well-understood for extracellular biomolecular interactions, we present here examples of, and discuss how, avidity and the underlying kinetics influences intracellular signaling processes

(S)Pinning down protein interactions by NMR

Protein molecules are highly diverse communication platforms and their interaction repertoire stretches from atoms over small molecules such as sugars and lipids to macromolecules. An important route to understanding molecular communication is to quantitatively describe their interactions. These types of analyses determine the amounts and proportions of individual constituents that participate in a reaction as well as their rates of reactions and their thermodynamics. Although many different methods are available, there is currently no single method able to quantitatively capture and describe all types of protein reactions, which can span orders of magnitudes in affinities, reaction rates, and lifetimes of states. As the more versatile technique, solution NMR spectroscopy offers a remarkable catalogue of methods that can be successfully applied to the quantitative as well as qualitative descriptions of protein interactions. In this review we provide an easy‐access approach to NMR for the non‐NMR specialist and describe how and when solution state NMR spectroscopy is the method of choice for addressing protein ligand interaction. We describe very briefly the theoretical background and illustrate simple protein–ligand interactions as well as typical strategies for measuring binding constants using NMR spectroscopy. Finally, this review provides examples of caveats of the method as well as the options to improve the outcome of an NMR analysis of a protein interaction reaction

Super-resolution microscopy reveals functional organization of dopamine transporters into cholesterol and neuronal activity-dependent nanodomains

The dopamine transporter (DAT) has a crucial role in the regulation of neurotransmission. Here, the authors use super-resolution imaging to show that DAT clusters into cholesterol-dependent membrane regions that are reversibly regulated by ionotropic glutamate receptors activation

A high-affinity, bivalent PDZ domain inhibitor complexes PICK1 to alleviate neuropathic pain

Maladaptive plasticity involving increased expression of AMPA‐type glutamate receptors is involved in several pathologies, including neuropathic pain, but direct inhibition of AMPARs is associated with side effects. As an alternative, we developed a cell‐permeable, high‐affinity (~2 nM) peptide inhibitor, Tat‐P$_4$‐(C5)$_2$, of the PDZ domain protein PICK1 to interfere with increased AMPAR expression. The affinity is obtained partly from the Tat peptide and partly from the bivalency of the PDZ motif, engaging PDZ domains from two separate PICK1 dimers to form a tetrameric complex. Bivalent Tat‐P$_4$‐(C5)$_2$ disrupts PICK1 interaction with membrane proteins on supported cell membrane sheets and reduce the interaction of AMPARs with PICK1 and AMPA‐receptor surface expression in vivo. Moreover, Tat‐P$_4$‐(C5)$_2$ administration reduces spinal cord transmission and alleviates mechanical hyperalgesia in the spared nerve injury model of neuropathic pain. Taken together, our data reveal Tat‐P$_4$‐(C5)$_2$ as a novel promising lead for neuropathic pain treatment and expand the therapeutic potential of bivalent inhibitors to non‐tandem protein–protein interaction domains

Membrane Binding and Modulation of the PDZ Domain of PICK1

Scaffolding proteins serve to assemble protein complexes in dynamic processes by means of specific protein-protein and protein-lipid binding domains. Many of these domains bind either proteins or lipids exclusively; however, it has become increasingly evident that certain domains are capable of binding both. Especially, many PDZ domains, which are highly abundant protein-protein binding domains, bind lipids and membranes. Here we provide an overview of recent large-scale studies trying to generalize and rationalize the binding patterns as well as specificity of PDZ domains towards membrane lipids. Moreover, we review how these PDZ-membrane interactions are regulated in the case of the synaptic scaffolding protein PICK1 and how this might affect cellular localization and function