Β-hairpin Folding Research Articles

Engineering Enhanced Protein Stability through β‐Turn Optimization: Insights for the Design of Stable Peptide β‐Hairpin Systems

One good turn: β-Turns are an important component of protein structure and nucleation sites for β-hairpin folding. With ubiquitin as a “host” system (see structure), protein engineering methods have been used to determine the energetic contribution of type I′ β-turns to the stability of proteins, with a view to optimizing the rational design of model peptide systems.

Combinatorial Pattern Discovery Approach for the Folding Trajectory Analysis of a β-Hairpin

The study of protein folding mechanisms continues to be one of the most challenging problems in computational biology. Currently, the protein folding mechanism is often characterized by calculating the free energy landscape versus various reaction coordinates, such as the fraction of native contacts, the radius of gyration, RMSD from the native structure, and so on. In this paper, we present a combinatorial pattern discovery approach toward understanding the global state changes during the folding process. This is a first step toward an unsupervised (and perhaps eventually automated) approach toward identification of global states. The approach is based on computing biclusters (or patterned clusters)—each cluster is a combination of various reaction coordinates, and its signature pattern facilitates the computation of the Z-score for the cluster. For this discovery process, we present an algorithm of time complexity c∈RO((N + nm) log n), where N is the size of the output patterns and (n × m) is the size of the input with n time frames and m reaction coordinates. To date, this is the best time complexity for this problem. We next apply this to a β-hairpin folding trajectory and demonstrate that this approach extracts crucial information about protein folding intermediate states and mechanism. We make three observations about the approach: (1) The method recovers states previously obtained by visually analyzing free energy surfaces. (2) It also succeeds in extracting meaningful patterns and structures that had been overlooked in previous works, which provides a better understanding of the folding mechanism of the β-hairpin. These new patterns also interconnect various states in existing free energy surfaces versus different reaction coordinates. (3) The approach does not require calculating the free energy values, yet it offers an analysis comparable to, and sometimes better than, the methods that use free energy landscapes, thus validating the choice of reaction coordinates. (An abstract version of this work was presented at the 2005 Asia Pacific Bioinformatics Conference [1].)

Kinetic Pathways of β-Hairpin (Un)folding in Explicit Solvent

We examine the dynamical (un)folding pathways of the C-terminal β-hairpin of protein G-B1 at room temperature in explicit solvent, by employing transition path sampling algorithms. The path ensembles contain information on the folding kinetics, including solvent motion. We determine the transition state ensembles for the two main transitions: 1), the hydrophobic collapse; and 2), the backbone hydrogen bond formation. In both cases the transition state ensembles are characterized by a layer ( 1) or a strip ( 2) of water molecules in between the two hairpin strands, supporting the hypothesis of the solvent as lubricant in the folding process. The transition state ensembles do not correspond with saddle points in the equilibrium free-energy landscapes. The kinetic pathways are thus not completely determined by the free-energy landscape. This phenomenon can occur if the order parameters obey different timescales. Using the transition interface sampling technique, we calculate the rate constants for (un)folding and find them in reasonable agreement with experiments, thus supporting the validation of using all-atom force fields to study protein folding.

Understanding the key factors that control the rate of β-hairpin folding

Both turn sequence and interstrand hydrophobic side-chain-side-chain interaction have been suggested to be important determinants of beta-hairpin stability. However, their roles in controlling the folding dynamics of beta-hairpins have not been clearly determined. Herein, we investigated the structural stability and folding kinetics of a series of tryptophan zippers by static IR and CD spectroscopies and the IR temperature jump method. Our results support a beta-hairpin folding mechanism wherein the rate-limiting event corresponds to the formation of the turn. We find that the logarithm of the folding rate depends linearly on the entropic change associated with the turn formation, where faster folding correlates with lower entropic cost. Moreover, a stronger turn-promoting sequence increases the stability of a beta-hairpin primarily by increasing its folding rate, whereas a stronger hydrophobic cluster increases the stability of a beta-hairpin primarily by decreasing its unfolding rate.

Comment on “Can a Continuum Solvent Model Reproduce the Free Energy Landscape of a β-Hairpin Folding in Water?” The Poisson−Boltzmann Equation

ADVERTISEMENT RETURN TO ISSUEPREVCommentComment on “Can a Continuum Solvent Model Reproduce the Free Energy Landscape of a β-Hairpin Folding in Water?” The Poisson−Boltzmann EquationRuhong Zhou, Goran Krilov, and B. J. BerneView Author Information Computational Biology Center, IBM Thomas J. Watson Research Center, 1101 Kitchawan Road, Yorktown Heights, New York 10598, and Department of Chemistry, Columbia University, New York, New York 10027 Cite this: J. Phys. Chem. B 2004, 108, 22, 7528–7530Publication Date (Web):May 4, 2004Publication History Received11 December 2003Revised18 February 2004Published online4 May 2004Published inissue 1 June 2004https://doi.org/10.1021/jp037812cCopyright © 2004 American Chemical SocietyRIGHTS & PERMISSIONSArticle Views222Altmetric-Citations26LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (159 KB) Get e-AlertsSUBJECTS:Chemical structure,Free energy,Hydrophobicity,Molecular mechanics,Solvents Get e-Alerts

Beta-hairpin folding and stability: molecular dynamics simulations of designed peptides in aqueous solution.

The structural properties of a 10-residue and a 15-residue peptide in aqueous solution were investigated by molecular dynamics simulation. The two designed peptides, SYINSDGTWT and SESYINSDGTWTVTE, had been studied previously by NMR at 278 K and the resulting model structures were classified as 3:5 beta-hairpins with a type I + G1 beta-bulge turn. In simulations at 278 K, starting from the NMR model structure, the 3:5 beta-hairpin conformers proved to be stable over the time period evaluated (30 ns). Starting from an extended conformation, simulations of the decapeptide at 278 K, 323 K and 353 K were also performed to study folding. Over the relatively short time scales explored (30 ns at 278 K and 323 K, 56 ns at 353 K), folding to the 3:5 beta-hairpin could only be observed at 353 K. At this temperature, the collapse to beta-hairpin-like conformations is very fast. The conformational space accessible to the peptide is entirely dominated by loop structures with different degrees of beta-hairpin character. The transitions between different types of ordered loops and beta-hairpins occur through two unstructured loop conformations stabilized by a single side-chain interaction between Tyr2 and Trp9, which facilitates the changes of the hydrogen-bond register. In agreement with previous experimental results, beta-hairpin formation is initially driven by the bending propensity of the turn segment. Nevertheless, the fine organization of the turn region appears to be a late event in the folding process.

β-Hairpin Folding Mechanism of a Nine-Residue Peptide Revealed from Molecular Dynamics Simulations in Explicit Water

The β-hairpin fold mechanism of a nine-residue peptide, which is modified from the β-hairpin of α-amylase inhibitor tendamistat (residues 15–23), is studied through direct folding simulations in explicit water at native folding conditions. Three 300-nanosecond self-guided molecular dynamics (SGMD) simulations have revealed a series of β-hairpin folding events. During these simulations, the peptide folds repeatedly into a major cluster of β-hairpin structures, which agree well with nuclear magnetic resonance experimental observations. This major cluster is found to have the minimum conformational free energy among all sampled conformations. This peptide also folds into many other β-hairpin structures, which represent some local free energy minimum states. In the unfolded state, the N-terminal residues of the peptide, Tyr-1, Gln-2, and Asn-3, have a confined conformational distribution. This confinement makes β-hairpin the only energetically favored structure to fold. The unfolded state of this peptide is populated with conformations with non-native intrapeptide interactions. This peptide goes through fully hydrated conformations to eliminate non-native interactions before folding into a β-hairpin. The folding of a β-hairpin starts with side-chain interactions, which bring two strands together to form interstrand hydrogen bonds. The unfolding of the β-hairpin is not simply the reverse of the folding process. Comparing unfolding simulations using MD and SGMD methods demonstrate that SGMD simulations can qualitatively reproduce the kinetics of the peptide system.

Folding Dynamics of β-Hairpins: Molecular Dynamics Simulations

We have studied the folding mechanism of β-hairpins from proteins of 1GB1, 3AIT and 1A2P by unfolding simulations at high temperatures. The analysis of trajectories obtained from molecular dynamics simulations in explicit aqueous solution suggests that the three β-hairpin structures follow different mechanism of folding. The results of unfolding simulations showed that the positions of the hydrophobic core residues influence the folding dynamics. We discussed the characteristics of different mechanisms of β-hairpin folding based on the hydrogen-bond-centric and the hydrophobic-centric models.

Can a continuum solvent model reproduce the free energy landscape of a beta -hairpin folding in water?

The folding free energy landscape of the C-terminal beta-hairpin of protein G is explored using the surface-generalized Born (SGB) implicit solvent model, and the results are compared with the landscape from an earlier study with explicit solvent model. The OPLSAA force field is used for the beta-hairpin in both implicit and explicit solvent simulations, and the conformational space sampling is carried out with a highly parallel replica-exchange method. Surprisingly, we find from exhaustive conformation space sampling that the free energy landscape from the implicit solvent model is quite different from that of the explicit solvent model. In the implicit solvent model some nonnative states are heavily overweighted, and more importantly, the lowest free energy state is no longer the native beta-strand structure. An overly strong salt-bridge effect between charged residues (E42, D46, D47, E56, and K50) is found to be responsible for this behavior in the implicit solvent model. Despite this, we find that the OPLSAA/SGB energies of all the nonnative structures are higher than that of the native structure; thus the OPLSAA/SGB energy is still a good scoring function for structure prediction for this beta-hairpin. Furthermore, the beta-hairpin population at 282 K is found to be less than 40% from the implicit solvent model, which is much smaller than the 72% from the explicit solvent model and approximately equal 80% from experiment. On the other hand, both implicit and explicit solvent simulations with the OPLSAA force field exhibit no meaningful helical content during the folding process, which is in contrast to some very recent studies using other force fields.

Simulations of beta-hairpin folding confined to spherical pores using distributed computing.

We report the thermodynamics and kinetics of an off-lattice Go model beta-hairpin from Ig-binding protein confined to an inert spherical pore. Confinement enhances the stability of the hairpin due to the decrease in the entropy of the unfolded state. Compared with their values in the bulk, the rates of hairpin formation increase in the spherical pore. Surprisingly, the dependence of the rates on the pore radius, R(s), is nonmonotonic. The rates reach a maximum at R(s)/R(g,N)(b) approximately equal to 1.5, where R(g,N)(b) is the radius of gyration of the folded beta-hairpin in the bulk. The denatured state ensemble of the encapsulated beta-hairpin is highly structured even at substantially elevated temperatures. Remarkably, a profound effect of confinement is evident even when the beta-hairpin occupies less than a 10th of the sphere volume. Our calculations show that the emergence of substantial structure in the denatured state of proteins in inert pores is a consequence of confinement. In contrast, the structure of the bulk denatured state ensemble depends dramatically on the extent of denaturation.

Folding of β-Hairpins

Identification of the factors governing the formation of β-structure independently of the rest of the protein is important for understanding the folding process of protein into a unique native structure. It has been shown that some β-hairpins can fold autonomously into native-like structures, either in aqueous solution or in the presence of an organic co-solvent. Our aim is to review recent theoretical and experimental studies of folding of β-structures.

PII: S0969-2126(01)00700-6

PII: S0969-2126(01)00700-6

Understanding β-Hairpin Formation by Molecular Dynamics Simulations of Unfolding

We have studied the mechanism of formation of a 16-residue β-hairpin from the protein GB1 using molecular dynamics simulations in an aqueous environment. The analysis of unfolding trajectories at high temperatures suggests a refolding pathway consisting of several transient intermediates. The changes in the interaction energies of residues are related with the structural changes during the unfolding of the hairpin. The electrostatic energies of the residues in the turn region are found to be responsible for the transition between the folded state and the hydrophobic core state. The van der Waals interaction energies of the residues in the hydrophobic core reflect the behavior of the radius of gyration of the core region. We have examined the opposing influences of the protein-protein (PP) energy, which favors the native state, and the protein-solvent (PS) energy, which favors unfolding, in the formation of the β-hairpin structure. It is found that the behavior of the electrostatic components of PP and PS energies reflects the structural changes associated with the loss of backbone hydrogen bonding. Relative changes in the PP and PS van der Waals interactions are related with the disruption of the hydrophobic core of a protein. The results of the simulations support the hydrophobic collapse mechanism of β-hairpin folding.

β-hairpin folding simulations in atomistic detail using an implicit solvent model

We have used distributed computing techniques and a supercluster of thousands of computer processors to study folding of the C-terminal β-hairpin from protein G in atomistic detail using the GB/SA implicit solvent model at 300 K. We have simulated a total of nearly 38 μs of folding time and obtained eight complete and independent folding trajectories. Starting from an extended state, we observe relaxation to an unfolded state characterized by non-specific, temporary hydrogen bonding. This is followed by the appearance of interactions between hydrophobic residues that stabilize a bent intermediate. Final formation of the complete hydrophobic core occurs cooperatively at the same time that the final hydrogen bonding pattern appears. The folded hairpin structures we observe all contain a closely packed hydrophobic core and proper β-sheet backbone dihedral angles, but they differ in backbone hydrogen bonding pattern. We show that this is consistent with the existing experimental data on the hairpin alone in solution. Our analysis also reveals short-lived semi-helical intermediates which define a thermodynamic trap. Our results are consistent with a three-state mechanism with a single rate-limiting step in which a varying final hydrogen bond pattern is apparent, and semi-helical off-pathway intermediates may appear early in the folding process. We include details of the ensemble dynamics methodology and a discussion of our achievements using this new computational device for studying dynamics at the atomic level.

β-hairpin folds by molecular dynamics simulations

Our molecular dynamics simulations of a distal hairpin from the src SH3 domain in explicit water demonstrate that the folding of a β-hairpin from the unfolded conformation can occur on an accessible time-scale and the AMBER C96 force field is appropriate for studying β-hairpin folding. The present study reinforces the experimental results that a proper turn conformation of the distal hairpin that is determined by the tertiary interactions is important for the folding of this hairpin to the native structure. Insight into the folding mechanism of β-hairpins is obtained from an analysis of the trajectories.

Role of side-chains in the cooperative beta-hairpin folding of the short C-terminal fragment derived from streptococcal protein G.

A short C-terminal fragment of immunoglobulin-binding domain of streptococcal protein G is known to form nativelike beta-hairpin at physiological conditions. To understand the cooperative folding of the short peptide, eight Ala-substituted mutants of the fragment were investigated with respect to their structural stabilities by analyzing temperature dependence of NMR signals. On comparison of the obtained thermodynamic parameters, we found that the nonpolar residues Tyr45 and Phe52 and the polar residues Asp46 and Thr49 are crucial for the beta-hairpin folding. The results suggest a strong interaction between the nonpolar side chains that participates in a putative hydrophobic cluster and that the polar side chains form a fairly rigid conformation around the loop (46-51). We also investigated the complex formation of the mutants with N-terminal fragment at the variety of temperature to get their thermal unfolding profiles and found that the mutations on the residues Asp46 and Thr49 largely destabilized the complexes, while substitution of Asp47 slightly stabilized the complex. From these results, we deduced that both the hydrophobic cluster formation and the rigidity of the loop (46-51) cooperatively stabilize the beta-hairpin structure of the fragment. These interactions which form a stable beta-hairpin may be the initial structural scaffold which is important in the early folding events of the whole domain.

β-hairpin stability and folding: molecular dynamics studies of the first β-hairpin of tendamistat

The stability and (un)folding of the 19-residue peptide, SCVTLYQSWRYSQADNGCA, corresponding to the first β-hairpin (residues 10 to 28) of the α-amylase inhibitor tendamistat (PDB entry 3AIT) has been studied by molecular dynamics simulations in explicit water under periodic boundary conditions at several temperatures (300 K, 360 K and 400 K), starting from various conformations for simulation lengths, ranging from 10 to 30 ns. Comparison of trajectories of the reduced and oxidized native peptides reveals the importance of the disulphide bridge closing the β-hairpin in maintaining a proper turn conformation, thereby insuring a proper side-chain arrangement of the conserved turn residues. This allows rationalization of the conservation of those cysteine residues among the family of α-amylase inhibitors. High temperature simulations starting from widely different initial configurations (native β-hairpin, α and left-handed helical and extended conformations) begin sampling similar regions of the conformational space within tens of nanoseconds, and both native and non-native β-hairpin conformations are recovered. Transitions between conformational clusters are accompanied by an increase in energy fluctuations, which is consistent with the increase in heat capacity measured experimentally upon protein folding. The folding events observed in the various simulations support a model for β-hairpin formation in which the turn is formed first, followed by hydrogen bond formation closing the hairpin, and subsequent stabilization by side-chain hydrophobic interactions.

Monte Carlo Simulations of β-Hairpin Folding at Constant Temperature

Monte Carlo simulations were applied to β-hairpin folding of a valine-based peptide. Two valine residues in the middle of the peptide were substituted with glycine, to serve as turn residues. Unlike lattice model simulations, structure prediction methods, and unfolding simulations, our simulations used an atom-based model, constant temperature (274 K), and non- β-hairpin initial conformations. Based on the concept of solvent reference, the effective energy function simplified the solvent calculation and overcame the multiple minima problem. Driven by the hydrophobic interaction, the peptide first folded into a compact U-shaped conformation with a central turn, in analogy to the initial collapse with simultaneous nucleation in protein folding. The peptide units in the U-shaped conformation then reoriented, gradually forming hydrogen bonds in the β-hairpin pattern from the β-turn to the ends of the strands. With the same energy function, an alanine-based peptide folded into helix-dominated structures. The basic structure types ( α-helix or β-hairpin) that formed during the simulations depended upon the amino acid sequence. Compared with helix, β-hairpin folding is driven mainly by the hydrophobic interaction. Hydrogen bonding is necessary to maintain the ordered secondary structure.

Proteolytic refolding of the HIV-1 capsid protein amino-terminus facilitates viral core assembly.

After budding, the human immunodeficiency virus (HIV) must 'mature' into an infectious viral particle. Viral maturation requires proteolytic processing of the Gag polyprotein at the matrix-capsid junction, which liberates the capsid (CA) domain to condense from the spherical protein coat of the immature virus into the conical core of the mature virus. We propose that upon proteolysis, the amino-terminal end of the capsid refolds into a beta-hairpin/helix structure that is stabilized by formation of a salt bridge between the processed amino-terminus (Pro1) and a highly conserved aspartate residue (Asp51). The refolded amino-terminus then creates a new CA-CA interface that is essential for assembling the condensed conical core. Consistent with this model, we found that recombinant capsid proteins with as few as four matrix residues fused to their amino-termini formed spheres in vitro, but that removing these residues refolded the capsid amino-terminus and redirected protein assembly from spheres to cylinders. Moreover, point mutations throughout the putative CA-CA interface blocked capsid assembly in vitro, core assembly in vivo and viral infectivity. Disruption of the conserved amino-terminal capsid salt bridge also abolished the infectivity of Moloney murine leukemia viral particles, suggesting that lenti- and oncoviruses mature via analogous pathways.

NMR evidence for the nucleation of a β-hairpin peptide conformation in water by an Asn-Gly type I′ β-turn sequence

The contribution of the β-turn sequence to the folding and stability of a peptide β-hairpin in water has been analysed from studies of a truncated peptide lacking one β-strand and hence the majority of the interstrand hydrophobic interactions; NMR analysis shows that the Asn-Gly type I′ β-turn conformation is significantly populated, suggesting that the intrinsic conformation preference of the turn sequence may play an important role in nucleating hairpin folding.