Transfer Free Energies of Test Proteins Into Crowded Protein Solutions Have Simple Dependence on Crowder Concentration.

The effects of macromolecular crowding on the thermodynamic properties of test proteins are determined by the latter's transfer free energies from a dilute solution to a crowded solution. The transfer free energies in turn are determined by effective protein-crowder interactions. When these interactions are modeled at the all-atom level, the transfer free energies may defy simple predictions.

Designed Mutations Alter the Binding Pathways of an Intrinsically Disordered Protein.

Many cellular functions, including signaling and regulation, are carried out by intrinsically disordered proteins (IDPs) binding to their targets. Experimental and computational studies have now significantly advanced our understanding of these binding processes. In particular, IDPs that become structured upon binding typically follow a dock-and-coalesce mechanism, whereby the docking of one IDP segment initiates the process, followed by on-target coalescence of remaining IDP segments.

Atomistic Modeling of Intrinsically Disordered Proteins Under Polyethylene Glycol Crowding: Quantitative Comparison with Experimental Data and Implication of Protein-Crowder Attraction.

The malleability of intrinsically disordered proteins (IDPs) has generated great interest in understanding how their conformations respond to crowded cellular environments. Experiments can report gross properties such as fluorescence resonance energy transfer (FRET) efficiency but cannot resolve the conformational ensembles of IDPs and their interactions with macromolecular crowders. Computation can in principle provide the latter information but in practice has been hampered by the enormous expense for realistic modeling of IDPs and crowders and for sufficient conformational sampling.

A conserved glycine harboring disease-associated mutations permits NMDA receptor slow deactivation and high Ca permeability.

A variety of de novo and inherited missense mutations associated with neurological disorders are found in the NMDA receptor M4 transmembrane helices, which are peripheral to the pore domain in eukaryotic ionotropic glutamate receptors. Subsets of these mutations affect receptor gating with dramatic effects, including in one instance halting it, occurring at a conserved glycine near the extracellular end of M4. Functional experiments and molecular dynamic simulations of constructs with and without substitutions at this glycine indicate that it acts as a hinge, permitting the intracellular portion of the ion channel to laterally expand.

Temperature-induced collapse of a disordered peptide observed by three sampling methods in molecular dynamics simulations.

The conformational ensembles of a disordered peptide, polyglutamine Q15, over a wide temperature range were sampled using multiple replicates of conventional molecular dynamics (cMD) simulations as well as two enhanced sampling methods, temperature replica exchange (TREMD) and replica exchange with solute tempering (REST). The radius of gyration, asphericity, secondary structure, and hydrogen bonding patterns were used for the comparison of the sampling methods. Overall, the three sampling methods generated similar conformational ensembles, with progressive collapse at higher temperatures.

Why Do Disordered and Structured Proteins Behave Differently in Phase Separation?

Intracellular membraneless organelles and their myriad cellular functions have garnered tremendous recent interest. It is becoming well accepted that they form via liquid-liquid phase separation (LLPS) of protein mixtures (often including RNA), where the organelles correspond to a protein-rich droplet phase coexisting with a protein-poor bulk phase. The major protein components contain disordered regions and often also RNA-binding domains, and the disordered fragments on their own easily undergo LLPS.

Liquid-Liquid Phase Separation of Patchy Particles Illuminates Diverse Effects of Regulatory Components on Protein Droplet Formation.

Recently many cellular functions have been associated with membraneless organelles, or protein droplets, formed by liquid-liquid phase separation (LLPS). Proteins in these droplets often contain RNA-binding domains, but the effects of RNA on LLPS have been controversial. To gain better understanding on the roles of RNA and other macromolecular regulators, here we used Gibbs-ensemble simulations to determine phase diagrams of two-component patchy particles, as models for mixtures of proteins with regulatory components.

Electrostatic Interactions in Protein Structure, Folding, Binding, and Condensation.

Charged and polar groups, through forming ion pairs, hydrogen bonds, and other less specific electrostatic interactions, impart important properties to proteins. Modulation of the charges on the amino acids, e.g.

The dock-and-coalesce mechanism for the association of a WASP disordered region with the Cdc42 GTPase.

Intrinsically disordered proteins (IDPs) play key roles in signaling and regulation. Many IDPs undergo folding upon binding to their targets. We have proposed that coupled folding and binding of IDPs generally follow a dock-and-coalesce mechanism, whereby a segment of the IDP, through diffusion, docks to its cognate subsite and, subsequently, the remaining segments coalesce around their subsites.

Structural modeling for the open state of an NMDA receptor.

NMDA receptors are tetrameric ligand-gated ion channels that are crucial for neurodevelopment and higher order processes such as learning and memory, and have been implicated in numerous neurological disorders. The lack of a structure for the channel open state has greatly hampered the understanding of the normal gating process and mechanisms of disease-associated mutations. Here we report the structural modeling for the open state of an NMDA receptor.

Advancing NMDA Receptor Physiology by Integrating Multiple Approaches.

NMDA receptors (NMDARs) are ion channels activated by the excitatory neurotransmitter glutamate and are essential to all aspects of brain function, including learning and memory formation. Missense mutations distributed throughout NMDAR subunits have been associated with an array of neurological disorders. Recent structural, functional, and computational studies have generated many insights into the activation process connecting glutamate binding to ion-channel opening, which is central to NMDAR physiology and pathophysiology.

Gating Motions and Stationary Gating Properties of Ionotropic Glutamate Receptors: Computation Meets Electrophysiology.

Ionotropic glutamate receptors (iGluRs) are tetrameric ligand-gated ion channels essential to all aspects of brain function, including higher order processes such as learning and memory. For decades, electrophysiology was the primary means for characterizing the function of iGluRs and gaining mechanistic insight. Since the turn of the century, structures of isolated water-soluble domains and transmembrane-domain-containing constructs have provided the basis for formulating mechanistic hypotheses.

Protein folding, binding, and droplet formation in cell-like conditions.

The many bystander macromolecules in the crowded cellular environments present both steric repulsion and weak attraction to proteins undergoing folding or binding and hence impact the thermodynamic and kinetic properties of these processes. The weak but nonrandom binding with bystander macromolecules may facilitate subcellular localization and biological function. Weak binding also leads to the emergence of a protein-rich droplet phase, which has been implicated in regulating a variety of cellular functions.

Electrostatic effects on the folding stability of FKBP12.

The roles of electrostatic interactions in protein folding stability have been a matter of debate, largely due to the complexity in the theoretical treatment of these interactions. We have developed computational methods for calculating electrostatic effects on protein folding stability. To rigorously test and further refine these methods, here we carried out experimental studies into electrostatic effects on the folding stability of the human 12-kD FK506 binding protein (FKBP12).

Fast Method for Computing Chemical Potentials and Liquid-Liquid Phase Equilibria of Macromolecular Solutions.

Chemical potential is a fundamental property for determining thermodynamic equilibria involving exchange of molecules, such as between two phases of molecular systems. Previously, we developed the fast Fourier transform (FFT)-based method for Modeling Atomistic Protein-crowder interactions (FMAP) to calculate excess chemical potentials according to the Widom insertion. Intermolecular interaction energies were expressed as correlation functions and evaluated via FFT.

Challenges in structural approaches to cell modeling.

Computational modeling is essential for structural characterization of biomolecular mechanisms across the broad spectrum of scales. Adequate understanding of biomolecular mechanisms inherently involves our ability to model them. Structural modeling of individual biomolecules and their interactions has been rapidly progressing.

Prediction of homoprotein and heteroprotein complexes by protein docking and template-based modeling: A CASP-CAPRI experiment.

We present the results for CAPRI Round 30, the first joint CASP-CAPRI experiment, which brought together experts from the protein structure prediction and protein-protein docking communities. The Round comprised 25 targets from amongst those submitted for the CASP11 prediction experiment of 2014. The targets included mostly homodimers, a few homotetramers, and two heterodimers, and comprised protein chains that could readily be modeled using templates from the Protein Data Bank.

Mechanism and rate constants of the Cdc42 GTPase binding with intrinsically disordered effectors.

Intrinsically disordered proteins (IDPs) are often involved in signaling and regulatory functions, through binding to cellular targets. Many IDPs undergo disorder-to-order transitions upon binding. Both the binding mechanisms and the magnitudes of the binding rate constants can have functional importance.

Protein Allostery and Conformational Dynamics.

The functions of many proteins are regulated through allostery, whereby effector binding at a distal site changes the functional activity (e.g., substrate binding affinity or catalytic efficiency) at the active site.

The Transmembrane Domain Mediates Tetramerization of α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid (AMPA) Receptors.

AMPA receptors (AMPARs) mediate fast excitatory neurotransmission in the central nervous system. Functional AMPARs are tetrameric complexes with a highly modular structure, consisting of four evolutionarily distinct structural domains: an amino-terminal domain (ATD), a ligand-binding domain (LBD), a channel-forming transmembrane domain (TMD), and a carboxyl-terminal domain (CTD). Here we show that the isolated TMD of the GluA1 AMPAR is fully capable of tetramerization.

A Jump-from-Cavity Pyrophosphate Ion Release Assisted by a Key Lysine Residue in T7 RNA Polymerase Transcription Elongation.

Pyrophosphate ion (PPi) release during transcription elongation is a signature step in each nucleotide addition cycle. The kinetics and energetics of the process as well as how it proceeds with substantial conformational changes of the polymerase complex determine the mechano-chemical coupling mechanism of the transcription elongation. Here we investigated detailed dynamics of the PPi release process in a single-subunit RNA polymerase (RNAP) from bacteriophage T7, implementing all-atom molecular dynamics (MD) simulations.

Disorder-to-Order Transition of an Active-Site Loop Mediates the Allosteric Activation of Sortase A.

Intrinsically disordered proteins and intrinsically disordered regions are implicated in many biological functions and associated with many diseases, but their conformational characterizations are challenging. The disordered β6/β7 loop of Staphylococcus aureus sortase A is involved in the binding of both sorting signals and calcium. Calcium binding allosterically activates the enzyme, but the detailed mechanism has been unclear.

Dynamically Driven Protein Allostery Exhibits Disparate Responses for Fast and Slow Motions.

There is considerable interest in the dynamic aspect of allosteric action, and in a growing list of proteins allostery has been characterized as being mediated predominantly by a change in dynamics, not a transition in conformation. For considering conformational dynamics, a protein molecule can be simplified into a number of relatively rigid microdomains connected by joints, corresponding to, e.g.

Mechanism-Based Mathematical Model for Gating of Ionotropic Glutamate Receptors.

We present a mathematical model for ionotropic glutamate receptors (iGluR's) that is built on mechanistic understanding and yields a number of thermodynamic and kinetic properties of channel gating. iGluR's are ligand-gated ion channels responsible for the vast majority of fast excitatory neurotransmission in the central nervous system. The effects of agonist-induced closure of the ligand-binding domain (LBD) are transmitted to the transmembrane channel (TMC) via interdomain linkers.

Two pathways mediate interdomain allosteric regulation in pin1.

Allostery is an essential means for regulating biomolecular functions and provides unique opportunities for drug design, yet our ability to elucidate allosteric mechanisms remains limited. Here, based on extensive molecular dynamics simulations, we present an atomistic picture of the pathways mediating the allosteric regulation of the PPIase domain of Pin1 by its WW domain. Two pathways jointly propagate the action of substrate-WW binding to produce closure and rigidification of three PPIase catalytic-site loops.