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Journal ArticleDOI

Protein disulfide engineering.

TL;DR: Progress in disulfide engineering is reviewed, with an emphasis on the issue of stability and computational methods that facilitate engineering efforts.
About: This article is published in FEBS Letters.The article was published on 2014-01-21. It has received 205 citations till now.
Citations
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Journal ArticleDOI
TL;DR: What has been learned about the major forces stabilizing proteins since the late 1980s when site‐directed mutagenesis became possible is summarized.

284 citations


Cites background from "Protein disulfide engineering."

  • ...A comprehensive review of this subject was recently published in this journal [60]....

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Journal ArticleDOI
11 Jan 2018
TL;DR: The most relevant enzymes that are used for industrial applications are provided and the strategies that are adopted to enhance enzyme stability and/or activity are discussed, along with some of the most relevant achievements.
Abstract: The catalytic properties of some selected enzymes have long been exploited to carry out efficient and cost-effective bioconversions in a multitude of research and industrial sectors, such as food, health, cosmetics, agriculture, chemistry, energy, and others. Nonetheless, for several applications, naturally occurring enzymes are not considered to be viable options owing to their limited stability in the required working conditions. Over the years, the quest for novel enzymes with actual potential for biotechnological applications has involved various complementary approaches such as mining enzyme variants from organisms living in extreme conditions (extremophiles), mimicking evolution in the laboratory to develop more stable enzyme variants, and more recently, using rational, computer-assisted enzyme engineering strategies. In this review, we provide an overview of the most relevant enzymes that are used for industrial applications and we discuss the strategies that are adopted to enhance enzyme stability and/or activity, along with some of the most relevant achievements. In all living species, many different enzymes catalyze fundamental chemical reactions with high substrate specificity and rate enhancements. Besides specificity, enzymes also possess many other favorable properties, such as, for instance, cost-effectiveness, good stability under mild pH and temperature conditions, generally low toxicity levels, and ease of termination of activity. As efficient natural biocatalysts, enzymes provide great opportunities to carry out important chemical reactions in several research and industrial settings, ranging from food to pharmaceutical, cosmetic, agricultural, and other crucial economic sectors.

177 citations

Journal ArticleDOI
TL;DR: This review will focus on factors affecting protein stability on a somewhat practical level, particularly from the view of a protein crystallographer.
Abstract: Protein stability is a topic of major interest for the biotechnology, pharmaceutical and food industries, in addition to being a daily consideration for academic researchers studying proteins. An understanding of protein stability is essential for optimizing the expression, purification, formulation, storage and structural studies of proteins. In this review, discussion will focus on factors affecting protein stability, on a somewhat practical level, particularly from the view of a protein crystallographer. The differences between protein conformational stability and protein compositional stability will be discussed, along with a brief introduction to key methods useful for analyzing protein stability. Finally, tactics for addressing protein-stability issues during protein expression, purification and crystallization will be discussed.

170 citations


Cites methods from "Protein disulfide engineering."

  • ...Disulfide engineering is a well established method for stabilizing proteins and for studying and modifying protein function and dynamics (Dombkowski et al., 2014)....

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Journal ArticleDOI
TL;DR: The computational stability design methods have advanced over the past two decades starting from methods that selectively addressed only some aspects of marginal stability, such as thermodynamic, cellular, and evolutionary principles and mechanisms that underlie marginal stability as mentioned in this paper.
Abstract: Proteins are increasingly used in basic and applied biomedical research.Many proteins, however, are only marginally stable and can be expressed in limited amounts, thus hampering research and applications. Research has revealed the thermodynamic, cellular, and evolutionary principles and mechanisms that underlie marginal stability. With this growing understanding, computational stability design methods have advanced over the past two decades starting from methods that selectively addressed only some aspects of marginal stability. Current methods are more general and, by combining phylogenetic analysis with atomistic design, have shown drastic improvements in solubility, thermal stability, and aggregation resistance while maintaining the protein’s primary molecular activity. Stability design is opening the way to rational engineering of improved enzymes, therapeutics, and vaccines and to the application of protein design methodology to large proteins and molecular activities that have proven challenging in...

168 citations

Journal ArticleDOI
TL;DR: In this review different algorithms for the prediction of beneficial mutation sites to enhance protein stability are summarized and the advantages and disadvantages of FoldX are highlighted.
Abstract: Improving protein stability is an important goal for basic research as well as for clinical and industrial applications but no commonly accepted and widely used strategy for efficient engineering is known. Beside random approaches like error prone PCR or physical techniques to stabilize proteins, e.g. by immobilization, in silico approaches are gaining more attention to apply target-oriented mutagenesis. In this review different algorithms for the prediction of beneficial mutation sites to enhance protein stability are summarized and the advantages and disadvantages of FoldX are highlighted. The question whether the prediction of mutation sites by the algorithm FoldX is more accurate than random based approaches is addressed.

141 citations

References
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Book
01 Oct 1986
TL;DR: This paper discusses the physical properties of polypeptides, the structure of which has been determined Crystallographically to High Resolution and its role in the biosynthesis of Proteins.
Abstract: Chemical Properties of Polypeptides Biosynthesis of Proteins Evolutionary and Genetic Origins of Protein Sequences Physical Interactions that Determine the Properties of Proteins Conformational Properties of Polypeptide Chains The Folded Conformations of Globular Proteins Proteins in Solution and in Membranes Interactions with Other Molecules Enzyme Catalysis Degradation Appendix: References to Protein Structures Determined Crystallographically to High Resolution

4,285 citations

Journal ArticleDOI
15 Oct 2010-Science
TL;DR: Simulation of the folding of a WW domain showed a well-defined folding pathway and simulation of the dynamics of bovine pancreatic trypsin inhibitor showed interconversion between distinct conformational states.
Abstract: Molecular dynamics (MD) simulations are widely used to study protein motions at an atomic level of detail, but they have been limited to time scales shorter than those of many biologically critical conformational changes. We examined two fundamental processes in protein dynamics—protein folding and conformational change within the folded state—by means of extremely long all-atom MD simulations conducted on a special-purpose machine. Equilibrium simulations of a WW protein domain captured multiple folding and unfolding events that consistently follow a well-defined folding pathway; separate simulations of the protein’s constituent substructures shed light on possible determinants of this pathway. A 1-millisecond simulation of the folded protein BPTI reveals a small number of structurally distinct conformational states whose reversible interconversion is slower than local relaxations within those states by a factor of more than 1000.

1,650 citations


"Protein disulfide engineering." refers background in this paper

  • ...The total time simulated for a protein is typically in the nanoseconds but can extend into microseconds, and recently the millisecond range, for small proteins modeled with advanced hardware [54]....

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Journal ArticleDOI
TL;DR: This review offers an outline of the origin of molecular dynamics simulation for protein systems and how it has developed into a robust and trusted tool, and covers more recent advances in theory and an illustrative selection of practical studies in which it played a central role.
Abstract: The term molecular mechanics (MM) refers to the use of simple potential-energy functions (e.g., harmonic oscillator or Coulombic potentials) to model molecular systems. Molecular mechanics approaches are widely applied in molecular structure refinement, molecular dynamics (MD) simulations, Monte Carlo (MC) simulations, and ligand-docking simulations. Typically, molecular mechanics models consist of spherical atoms connected by springs which represent bonds. Internal forces experienced in the model structure are described using simple mathematical functions. For example, Hooke’s law is commonly used to describe bonded interactions, and the nonbonded atoms might be treated as inelastic hard spheres or may interact according to a Lennard-Jones potential. Using these simple models, a molecular dynamics simulation numerically solves Newton’s equations of motion, thus allowing structural fluctuations to be observed with respect to time. Dynamic simulation methods are widely used to obtain information on the time evolution of conformations of proteins and other biological macromolecules1–4 and also kinetic and thermodynamic information. Simulations can provide fine detail concerning the motions of individual particles as a function of time. They can be utilized to quantify the properties of a system at a precision and on a time scale that is otherwise inaccessible, and simulation is, therefore, a valuable tool in extending our understanding of model systems. Theoretical consideration of a system additionally allows one to investigate the specific contributions to a property through “computational alchemy”,5 that is, modifying the simulation in a way that is nonphysical but nonetheless allows a model’s characteristics to be probed. One particular example is the artificial conversion of the energy function from that representing one system to that of another during a simulation. This is an important technique in free-energy calculations.6 Thus, molecular dynamics simulations, along with a range of complementary computational approaches, have become valuable tools for investigating the basis of protein structure and function. This review offers an outline of the origin of molecular dynamics simulation for protein systems and how it has developed into a robust and trusted tool. This review then covers more recent advances in theory and an illustrative selection of practical studies in which it played a central role. The range of studies in which MD has played a considerable or pivotal role is immense, and this review cannot do justice to them; MD simulations of biomedical importance were recently reviewed.4 Particular emphasis will be placed on the study of dynamic aspects of protein recognition, an area where molecular dynamics has scope to provide broad and far-ranging insights. This review concludes with a brief discussion of the future potential offered to advancement of the biological and biochemical sciences and the remaining issues that must be overcome to allow the full extent of this potential to be realized. 1.1. Historical Background MD methods were originally conceived within the theoretical physics community during the 1950s. In 1957, Alder and Wainwright7 performed the earliest MD simulation using the so-called hard-sphere model, in which the atoms interacted only through perfect collisions. Rahman8 subsequently applied a smooth, continuous potential to mimic real atomic interactions. During the 1970s, as computers became more widespread, MD simulations were developed for more complex systems, culminating in 1976 with the first simulation of a protein9,10 using an empirical energy function constructed using physics-based first-principles assumptions. MD simulations are now widely and routinely applied and especially popular in the fields of materials science11,12 and biophysics. As will be discussed later in this review, a variety of experimental conditions may be simulated with modern theories and algorithms. The initial simulations only considered single molecules in vacuo. Over time, more realistic or at least biologically relevant simulations could be performed. This trend is continuing today. The initial protein MD simulation, of the small bovine pancreatic trypsin inhibitor (BPTI), covered only 9.2 ps of simulation time. Modern simulations routinely have so-called equilibration periods much longer than that, and production simulations of tens of nanoseconds are routine, with the first microsecond MD simulation being reported in 1998.13 In addition, the original BPTI simulation included only about 500 atoms rather than the 104-106 atoms that are common today. While much of this advancement results from an immense increase in availability of computing power, major theoretical and methodological developments also contribute significantly. The number of publications regarding MD theory and application of MD to biological systems is growing at an extraordinary pace. A single review cannot do justice to the recent applications of MD. Using data from ISI Web of Science, the authors estimate that during 2005 at least 800 articles will be published that discuss molecular dynamics and proteins. The historical counts are shown in Figure 1. Open in a separate window Figure 1 Articles matching ISI Web of Science query “TS=(protein) AND TS=(molecular dynamics)”.

999 citations


"Protein disulfide engineering." refers background in this paper

  • ...A comprehensive review of MD for protein simulations is available in [55]....

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Journal ArticleDOI
18 Oct 1990-Nature
TL;DR: Molecular dynamics is also being used to determine protein structures from NMR, to refine protein X-ray crystal structures faster from poorer starting models, and to calculate the free energy changes resulting from mutations in proteins.
Abstract: Molecular dynamics—the science of simulating the motions of a system of particles—applied to biological macromolecules gives the fluctuations in the relative positions of the atoms in a protein or in DNA as a function of time. Knowledge of these motions provides insights into biological phenomena such as the role of flexibility in ligand binding and the rapid solvation of the electron transfer state in photosynthesis. Molecular dynamics is also being used to determine protein structures from NMR, to refine protein X-ray crystal structures faster from poorer starting models, and to calculate the free energy changes resulting from mutations in proteins.

910 citations


"Protein disulfide engineering." refers methods in this paper

  • ...MD is a computational technique used to simulate the motion of atoms in a molecule as a function of time [53]....

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Journal ArticleDOI
TL;DR: There is a strong preference for shorter connections, with half-cystines separated by less than 24 residues in 49% of all disulphides.

747 citations


"Protein disulfide engineering." refers background in this paper

  • ...The seminal work of Janet Thornton in 1981 characterized the features and bond geometry of disulfides by analyzing the atomic coordinates of 55 disulfide bonds that existed in protein structures available at the time [2]....

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