Microsecond simulations of the folding/unfolding thermodynamics of the Trp‐cage miniprotein

We study the unbiased folding/unfolding thermodynamics of the Trp‐cage miniprotein using detailed molecular dynamics simulations of an all‐atom model of the protein in explicit solvent using the Amberff99SB force field. Replica‐exchange molecular dynamics simulations are used to sample the protein ensembles over a broad range of temperatures covering the folded and unfolded states at two densities. The obtained ensembles are shown to reach equilibrium in the 1 μs/replica timescale. The total simulation time used in the calculations exceeds 100 μs. Ensemble averages of the fraction folded, pressure, and energy differences between the folded and unfolded states as a function of temperature are used to model the free energy of the folding transition, ΔG(P, T), over the whole region of temperatures and pressures sampled in the simulations. The ΔG(P, T) diagram describes an ellipse over the range of temperatures and pressures sampled, predicting that the system can undergo pressure‐induced unfolding and cold denaturation at low temperatures and high pressures, and unfolding at low pressures and high temperatures. The calculated free energy function exhibits remarkably good agreement with the experimental folding transition temperature (T_f = 321 K), free energy, and specific heat changes. However, changes in enthalpy and entropy are significantly different than the experimental values. We speculate that these differences may be due to the simplicity of the semiempirical force field used in the simulations and that more elaborate force fields may be required to describe appropriately the thermodynamics of proteins. Proteins 2010. © 2010 Wiley‐Liss, Inc.     

Complete characterization of the mutation landscape reveals the effect on amylin stability and amyloidogenicity

Type‐II diabetes is believed to be partially aggravated by the emergence of toxic amylin protein deposits in the extracellular space of the pancreas β‐cells. Amylin, the regulatory hormone that is co‐secreted with insulin, has been observed to misfold into toxic structures. Pramlintide, an FDA approved injectable amylin analog mutated at positions 25, 28, and 29 was therefore developed to create a more stable, soluble, less‐aggregating, and equipotent peptide that is used as an adjunctive therapy for diabetes. However, because Pramlintide is not ideal, researchers have been exploring other amylin analogs as therapeutic replacements. In this work, we assist the finding of optimal analogs by computationally revealing the mutational landscape of amylin. We computed the structure energies of all possible single‐point mutations and studied the effect they have on amylin stability and amyloidogenicity. Each of the 37 amylin residues was mutated in silico into the 19 canonical amino acids and an energy function computing the Lennard–Jones, Coulomb and solvation energy was used to analyze changes in stability. The mutation landscape identified amylin's conserved stable regions, residues that can be tweaked to further stabilize structure, regions that are susceptible to mutations, and mutations that are amyloidogenic. We used the single‐point mutational landscape data to generate estimations for higher‐order multiple‐point mutational landscapes and discovered millions of three‐point mutations that are more stable and less amyloidogenic than Pramlintide. The landscapes provided an explanation for the effect of the S20G and Q10R mutations on the onset of diabetes of the Chinese and Maori populations, respectively. Proteins 2015; 83:1014–1026. © 2015 Wiley Periodicals, Inc.     

The effect of specific proline residues on the kinetic stability of the triosephosphate isomerases of two trypanosomes

The effect of specific residues on the kinetic stability of two closely related triosephosphate isomerases (from Trypanosoma cruzi, TcTIM and Trypanosoma brucei, TbTIM) has been studied. Based on a comparison of their β‐turn occurrence, we engineered two chimerical enzymes where their super secondary β‐loop‐α motifs 2 ((βα)₂) were swapped. Differential scanning calorimetry (DSC) experiments showed that the (βα)₂ motif of TcTIM inserted into TbTIM (2Tc) increases the kinetic stability. On the other hand, the presence of the (βα)₂ motif of TbTIM inserted into TcTIM (2Tb) gave a chimerical protein difficult to purify in soluble form and with a significantly reduced kinetic stability. The comparison of the contact maps of the (βα)₂ of TbTIM and TcTIM showed differences in the contact pattern of residues 43 and 49. In TcTIM these residues are prolines, located at the N‐terminal of loop‐2 and the C‐terminal of α‐helix‐2. Twelve mutants were engineered involving residues 43 and 49 to study the effect over the unfolding activation energy barrier (E_A). A systematic analysis of DSC data showed a large decrease on the E_A of TcTIM (ΔE_A ranging from 468 to 678 kJ/mol) when the single and double proline mutations are present. The relevance of Pro43 to the kinetic stability is also revealed by mutation S43P, which increased the free energy of the transition state of TbTIM by 17.7 kJ/mol. Overall, the results indicate that protein kinetic stability can be severely affected by punctual mutations, disturbing the complex network of interactions that, in concerted action, determine protein stability. Proteins 2017; 85:571–579. © 2016 Wiley Periodicals, Inc.     

Folding thermodynamics of β‐hairpins studied by replica‐exchange molecular dynamics simulations

We study the differences in folding stability of β‐hairpin peptides, including GB1 hairpin and a point mutant GB1 K10G, as well as tryptophan zippers (TrpZips): TrpZip1, TrpZip2, TrpZip3‐1, and TrpZip4. By performing replica‐exchange molecular dynamics simulations with Amber03* force field (a modified version of Amber ff03) in explicit solvent, we observe ab initio folding of all the peptides except TrpZip3‐1, which is experimentally known to be the least stable among the peptides studied here. By calculating the free energies of unfolding of the peptides at room temperature and folding midpoint temperatures for thermal unfolding of peptides, we find that TrpZip4 and GB1 K10G peptides are the most stable β‐hairpins followed by TrpZip1, GB1, and TrpZip2 in the given order. Hence, the proposed K10G mutation of GB1 peptide results in enhanced stability compared to wild‐type GB1. An important goal of our study is to test whether simulations with Amber 03* model can reproduce experimentally predicted folding stability differences between these peptides. While the stabilities of GB1 and TrpZip1 yield close agreement with experiment, TrpZip2 is found to be less stable than predicted by experiment. However, as heterogenous folding of TrpZip2 may yield divergent thermodynamic parameters by different spectroscopic methods, mismatching of results with previous experimental values are not conclusive of model shortcomings. For most of the cases, molecular simulations with Amber03* can successfully reproduce experimentally known differences between the mutated peptides, further highlighting the predictive capabilities of current state‐of‐the‐art all‐atom protein force fields. Proteins 2015; 83:1307–1315. © 2015 Wiley Periodicals, Inc.     

Exploring additivity effects of double mutations on the binding affinity of protein‐protein complexes

Additivity in binding affinity of protein‐protein complexes refers to the change in free energy of binding (ΔΔG_bind) for double (or multiple) mutations which is approximately equal to the sum of their corresponding single mutation ΔΔG_bind values. In this study, we have explored the additivity effect of double mutants, which shows a linear relationship between the binding affinity of double and sum of single mutants with a correlation of 0.90. However, the comparison of ΔΔG_bind values showed a mean absolute deviation of 0.86 kcal/mol, and 25.6% of the double mutants show a deviation of more than 1 kcal/mol, which are identified as non‐additive. The additivity effects have been analyzed based on the influence of structural features such as accessible surface area, long range order, binding propensity change, surrounding hydrophobicity, flexibility, atomic contacts between the mutations and distance between the 2 mutations. We found that non‐additive mutations tend to be closer to each other and have more contacts. We have also used machine learning methods to discriminate additive and non‐additive mutations using structure‐based features, which showed the accuracies in the range of 0.77–0.92 for protein‐protein complexes belonging to different functions. Further, we have compared the additivity effects of protein stability along with binding affinity and explored the similarities and differences between them. The results obtained in this study provide insights into the effects of various structural features on binding affinity of double mutants, and will aid the development of accurate methods to predict the binding affinity of double mutants.     

Highly Efficient and Stable Perovskite Solar Cells Using a Dopant‐Free Inexpensive Small Molecule as the Hole‐Transporting Material

The hole transporting layer (HTL) plays an important role in realizing efficient and stable perovskite solar cells (PSCs). In spite of intensive research efforts toward the development of HTL materials, low‐cost, dopant‐free hole transporting materials that lead to efficient and stable PSCs remain elusive. Herein, a simple polycyclic heteroaromatic hydrocarbon‐based small molecule, 2,5,9,12‐tetra(tert‐butyl)diacenaphtho[1,2‐b:1′,2′‐d]thiophenen, as an efficient HTL material in PSCs is presented. This molecule is easy to synthesize and inexpensive. It is hydrophobic and exhibits excellent film‐forming properties on perovskites. It has unusually high hole mobility and a desirable highest occupied molecular orbital energy level, making it an ideal HTL material. PSCs fabricated using both the n‐i‐p planar and mesoscopic architectures with this compound as the HTL show efficiencies as high as 15.59% and 18.17%, respectively, with minimal hysteresis and high long term stability under ambient conditions.     

Osmolyte‐induced conformational changes in the Hsp90 molecular chaperone

Osmolytes are small molecules that play a central role in cellular homeostasis and the stress response by maintaining protein thermodynamic stability at controlled levels. The underlying physical chemistry that describes how different osmolytes impact folding free energy is well understood, however little is known about their influence on other crucial aspects of protein behavior, such as native‐state conformational changes. Here we investigate this issue with the Hsp90 molecular chaperone, a large dimeric protein that populates a complex conformational equilibrium. Using small angle X‐ray scattering we observe dramatic osmolyte‐dependent structural changes within the native ensemble. The degree to which different osmolytes affect the Hsp90 conformation strongly correlates with thermodynamic metrics of their influence on stability. This observation suggests that the well‐established osmolyte principles that govern stability also apply to large‐scale conformational changes, a proposition that is corroborated by structure‐based fitting of the scattering data, surface area comparisons and m‐value analysis. This approach shows how osmolytes affect a highly cooperative open/closed structural transition between two conformations that differ by a domain‐domain interaction. Hsp90 adopts an additional ligand‐specific conformation in the presence of ATP and we find that osmolytes do not significantly affect this conformational change. Together, these results extend the scope of osmolytes by suggesting that they can maintain protein conformational heterogeneity at controlled levels using similar underlying principles that allow them to maintain protein stability; however the relative impact of osmolytes on different structural states can vary significantly.     

Dopant‐Free Hole Transporting Polymers for High Efficiency,Environmentally Stable Perovskite Solar Cells

Over the past five years, a rapid progress in organometal‐halide perovskite solar cells has greatly influenced emerging solar energy science and technology. In perovksite solar cells, the overlying hole transporting material (HTM) is critical for achieving high power conversion efficiencies (PCEs) and for protecting the air‐sensitive perovskite active layer. This study reports the synthesis and implementation of a new polymeric HTM series based on semiconducting 4,8‐dithien‐2‐yl‐benzo[1,2‐d;4,5‐d′]bistriazole‐alt‐benzo[1,2‐b:4,5‐b′]dithiophenes (pBBTa‐BDTs), yielding high PCEs and environmentally‐stable perovskite cells. These intrinsic (dopant‐free) HTMs achieve a stabilized PCE of 12.3% in simple planar heterojunction cells—the highest value to date for a polymeric intrinsic HTM. This high performance is attributed to efficient hole extraction/collection (the most efficient pBBTa‐BDT is highly ordered and orients π‐face‐down on the perovskite surface) and balanced electron/hole transport. The smooth, conformal polymer coatings suppress aerobic perovskite film degradation, significantly enhancing the solar cell 85 °C/65% RH PCE stability versus typical molecular HTMs.     

Mice with endogenous TDP‐43 mutations exhibit gain of splicing function and characteristics of amyotrophic lateral sclerosis

TDP‐43 (encoded by the gene TARDBP) is an RNA binding protein central to the pathogenesis of amyotrophic lateral sclerosis (ALS). However, how TARDBP mutations trigger pathogenesis remains unknown. Here, we use novel mouse mutants carrying point mutations in endogenous Tardbp to dissect TDP‐43 function at physiological levels both in vitro and in vivo. Interestingly, we find that mutations within the C‐terminal domain of TDP‐43 lead to a gain of splicing function. Using two different strains, we are able to separate TDP‐43 loss‐ and gain‐of‐function effects. TDP‐43 gain‐of‐function effects in these mice reveal a novel category of splicing events controlled by TDP‐43, referred to as “skiptic” exons, in which skipping of constitutive exons causes changes in gene expression. In vivo, this gain‐of‐function mutation in endogenous Tardbp causes an adult‐onset neuromuscular phenotype accompanied by motor neuron loss and neurodegenerative changes. Furthermore, we have validated the splicing gain‐of‐function and skiptic exons in ALS patient‐derived cells. Our findings provide a novel pathogenic mechanism and highlight how TDP‐43 gain of function and loss of function affect RNA processing differently, suggesting they may act at different disease stages.     

Structural and functional analysis of perforin mutations in association with clinical data of familial hemophagocytic lymphohistiocytosis type 2 (FHL2) patients

Perforin plays a key role in the immune system via pore formation at the target cell membrane in the elimination of virus‐infected and transformed cells. A vast number of observed mutations in perforin impair this mechanism resulting in a rare but fatal disease, familial hemophagocytic lymphohistiocytosis type 2 (FHL2). Here we report a comprehensive in silico structural analysis of a collection of 76 missense perforin mutations based on a proposed pore model. In our model, perforin monomers oligomerize having cyclic symmetry in consistent with previously found experimental constraints yet having flexibility in the size of the pore and the number of monomers involved. Clusters of the mutations on the model map to three distinct functional regions of the perforin. Calculated stability (free energy) changes show that the mutations mainly destabilize the protein structure, interestingly however, A91V polymorphism, leads to a more stable one. Structural characteristics of mutations help explain the severe functional consequences on perforin deficient patients. Our study provides a structural approach to the mutation effects on the perforin oligomerization and impaired cytotoxic function in FHL2 patients.     

Critical biophysical properties in the Pseudomonas aeruginosa efflux gene regulator MexR are targeted by mutations conferring multidrug resistance

The self‐assembling MexA‐MexB‐OprM efflux pump system, encoded by the mexO operon, contributes to facile resistance of Pseudomonas aeruginosa by actively extruding multiple antimicrobials. MexR negatively regulates the mexO operon, comprising two adjacent MexR binding sites, and is as such highly targeted by mutations that confer multidrug resistance (MDR). To understand how MDR mutations impair MexR function, we studied MexR‐wt as well as a selected set of MDR single mutants distant from the proposed DNA‐binding helix. Although DNA affinity and MexA‐MexB‐OprM repression were both drastically impaired in the selected MexR‐MDR mutants, MexR‐wt bound its two binding sites in the mexO with high affinity as a dimer. In the MexR‐MDR mutants, secondary structure content and oligomerization properties were very similar to MexR‐wt despite their lack of DNA binding. Despite this, the MexR‐MDR mutants showed highly varying stabilities compared with MexR‐wt, suggesting disturbed critical interdomain contacts, because mutations in the DNA‐binding domains affected the stability of the dimer region and vice versa. Furthermore, significant ANS binding to MexR‐wt in both free and DNA‐bound states, together with increased ANS binding in all studied mutants, suggest that a hydrophobic cavity in the dimer region already shown to be involved in regulatory binding is enlarged by MDR mutations. Taken together, we propose that the biophysical MexR properties that are targeted by MDR mutations—stability, domain interactions, and internal hydrophobic surfaces—are also critical for the regulation of MexR DNA binding.     

High‐Performance and Stable Perovskite Solar Cells Based on Dopant‐Free Arylamine‐Substituted Copper(II) Phthalocyanine Hole‐Transporting Materials

A power conversion efficiency (PCE) as high as 19.7% is achieved using a novel, low‐cost, dopant‐free hole transport material (HTM) in mixed‐ion solution‐processed perovskite solar cells (PSCs). Following a rational molecular design strategy, arylamine‐substituted copper(II) phthalocyanine (CuPc) derivatives are selected as HTMs, reaching the highest PCE ever reported for PSCs employing dopant‐free HTMs. The intrinsic thermal and chemical properties of dopant‐free CuPcs result in PSCs with a long‐term stability outperforming that of the benchmark doped 2,2′,7,7′‐Tetrakis‐(N,N‐di‐p‐methoxyphenylamine)‐9,9′‐Spirobifluorene (Spiro‐OMeTAD)‐based devices. The combination of molecular modeling, synthesis, and full experimental characterization sheds light on the nanostructure and molecular aggregation of arylamine‐substituted CuPc compounds, providing a link between molecular structure and device properties. These results reveal the potential of engineering CuPc derivatives as dopant‐free HTMs to fabricate cost‐effective and highly efficient PSCs with long‐term stability, and pave the way to their commercial‐scale manufacturing. More generally, this case demonstrates how an integrated approach based on rational design and computational modeling can guide and anticipate the synthesis of new classes of materials to achieve specific functions in complex device structures.     

DynaMut2: Assessing changes in stability and flexibility upon single and multiple point missense mutations

Predicting the effect of missense variations on protein stability and dynamics is important for understanding their role in diseases, and the link between protein structure and function. Approaches to estimate these changes have been proposed, but most only consider single‐point missense variants and a static state of the protein, with those that incorporate dynamics are computationally expensive. Here we present DynaMut2, a web server that combines Normal Mode Analysis (NMA) methods to capture protein motion and our graph‐based signatures to represent the wildtype environment to investigate the effects of single and multiple point mutations on protein stability and dynamics. DynaMut2 was able to accurately predict the effects of missense mutations on protein stability, achieving Pearson's correlation of up to 0.72 (RMSE: 1.02 kcal/mol) on a single point and 0.64 (RMSE: 1.80 kcal/mol) on multiple‐point missense mutations across 10‐fold cross‐validation and independent blind tests. For single‐point mutations, DynaMut2 achieved comparable performance with other methods when predicting variations in Gibbs Free Energy (ΔΔG) and in melting temperature (ΔT_m). We anticipate our tool to be a valuable suite for the study of protein flexibility analysis and the study of the role of variants in disease. DynaMut2 is freely available as a web server and API at http://biosig.unimelb.edu.au/dynamut2 .     

Molecular dynamics simulations on interaction between bacterial proteins: Implication on pathogenic activities

We perform molecular dynamics simulation studies on interaction between bacterial proteins: an outer‐membrane protein STY3179 and a yfdX protein STY3178 of Salmonella Typhi. STY3179 has been found to be involved in bacterial adhesion and invasion. STY3178 is recently biophysically characterized. It is a soluble protein having antibiotic binding and chaperon activity capabilities. These two proteins co‐occur and are from neighboring gene in Salmonella Typhi‐occurrence of homologs of both STY3178 and STY3179 are identified in many Gram‐negative bacteria. We show using homology modeling, docking followed by molecular dynamics simulation that they can form a stable complex. STY3178 belongs to aqueous phase, while the beta barrel portion of STY3179 remains buried in DPPC bilayer with extra‐cellular loops exposed to water. To understand the molecular basis of interaction between STY3178 and STY3179, we compute the conformational thermodynamics which indicate that these two proteins interact through polar and acidic residues belonging to their interfacial region. Conformational thermodynamics results further reveal instability of certain residues in extra‐cellular loops of STY3179 upon complexation with STY3178 which is an indication for binding with host cell protein laminin.     

Development of Dopant‐Free Organic Hole Transporting Materials for Perovskite Solar Cells

There has been considerable progress over the last decade in development of the perovskite solar cells (PSCs), with reported performances now surpassing 25.2% power conversion efficiency. Both long‐term stability and component costs of PSCs remain to be addressed by the research community, using hole transporting materials (HTMs) such as 2,2′,7,7′‐tetrakis(N,N′‐di‐pmethoxyphenylamino)‐9,9′‐spirbiuorene(Spiro‐OMeTAD) and poly[bis(4‐phenyl)(2,4,6‐trimethylphenyl)amine] (PTAA). HTMs are essential for high‐performance PSC devices. Although effective, these materials require a relatively high degree of doping with additives to improve charge mobility and interlayer/substrate compatibility, introducing doping‐induced stability issues with these HTMs, and further, additional costs and experimental complexity associated with using these doped materials. This article reviews dopant‐free organic HTMs for PSCs, outlining reports of structures with promising properties toward achieving low‐cost, effective, and scalable materials for devices with long‐term stability. It summarizes recent literature reports on non‐doped, alternative, and more stable HTMs used in PSCs as essential components for high‐efficiency cells, categorizing HTMs as reported for different PSC architectures in addition to use of dopant‐free small molecular and polymeric HTMs. Finally, an outlook and critical assessment of dopant‐free organic HTMs toward commercial application and insight into the development of stable PSC devices is provided.     

Steric and thermodynamic limits of design for the incorporation of large unnatural amino acids in aminoacyl‐tRNA synthetase enzymes

Orthogonal aminoacyl‐tRNA synthetase/tRNA pairs from archaea have been evolved to facilitate site specific in vivo incorporation of unnatural amino acids into proteins in Escherichia coli. Using this approach, unnatural amino acids have been successfully incorporated with high translational efficiency and fidelity. In this study, CHARMM‐based molecular docking and free energy calculations were used to evaluate rational design of specific protein–ligand interactions for aminoacyl‐tRNA synthetases. A series of novel unnatural amino acid ligands were docked into the p‐benzoyl‐L ‐phenylalanine tRNA synthetase, which revealed that the binding pocket of the enzyme does not provide sufficient space for significantly larger ligands. Specific binding site residues were mutated to alanine to create additional space to accommodate larger target ligands, and then mutations were introduced to improve binding free energy. This approach was used to redesign binding sites for several different target ligands, which were then tested against the standard 20 amino acids to verify target specificity. Only the synthetase designed to bind Man‐α‐O‐Tyr was predicted to be sufficiently selective for the target ligand and also thermodynamically stable. Our study suggests that extensive redesign of the tRNA synthatase binding pocket for large bulky ligands may be quite thermodynamically unfavorable. Proteins 2010. © 2010 Wiley‐Liss, Inc.     

Cysteine‐free rop: A four‐helix bundle core mutant has wild‐type stability and structure but dramatically different unfolding kinetics

Cysteine residues can complicate the folding and storage of proteins due to improper formation of disulfide bonds or oxidation of residues that are natively reduced. Wild‐type Rop is a homodimeric four‐helix bundle protein and an important model for protein design in the understanding of protein stability, structure and folding kinetics. In the native state, Rop has two buried, reduced cysteine residues in its core, but these are prone to oxidation in destabilized variants, particularly upon extended storage. To circumvent this problem, we designed and characterized a Cys‐free variant of Rop, including solving the 2.3 Å X‐ray crystal structure. We show that the C38A C52V variant has similar structure, stability and in vivo activity to wild‐type Rop, but that it has dramatically faster unfolding kinetics like virtually every other mutant of Rop that has been characterized. This cysteine‐free Rop has already proven useful for studies on solution topology and on the relationship of core mutations to stability. It also suggests a general strategy for removal of pairs of Cys residues in proteins, both to make them more experimentally tractable and to improve their storage properties for therapeutic or industrial purposes.     

Metabolic systems maintain stable non‐equilibrium via thermodynamic buffering

Here, we analyze how the set of nucleotides in the cell is equilibrated and how this generates simple rules that help the cell to organize itself via maintenance of a stable non‐equilibrium state. A major mechanism operating to achieve this state is thermodynamic buffering via high activities of equilibrating enzymes such as adenylate kinase. Under stable non‐equilibrium, the ratios of free and Mg‐bound adenylates, Mg²⁺ and membrane potentials are interdependent and can be computed. The adenylate status is balanced with the levels of reduced and oxidized pyridine nucleotides through regulated uncoupling of the pyridine nucleotide pool from ATP production in mitochondria, and through oxidation of substrates non‐coupled to NAD⁺ reduction in peroxisomes. The set of adenylates and pyridine nucleotides constitutes a generalized cell energy status and determines rates of major metabolic fluxes. As the result, fluxes of energy and information become organized spatially and temporally, providing conditions for self‐maintenance of metabolism.     

pH‐selective mutagenesis of protein–protein interfaces: In silico design of therapeutic antibodies with prolonged half‐life

Understanding the effects of mutation on pH‐dependent protein binding affinity is important in protein design, especially in the area of protein therapeutics. We propose a novel method for fast in silico mutagenesis of protein–protein complexes to calculate the effect of mutation as a function of pH. The free energy differences between the wild type and mutants are evaluated from a molecular mechanics model, combined with calculations of the equilibria of proton binding. The predicted pH‐dependent energy profiles demonstrate excellent agreement with experimentally measured pH‐dependency of the effect of mutations on the dissociation constants for the complex of turkey ovomucoid third domain (OMTKY3) and proteinase B. The virtual scanning mutagenesis identifies all hotspots responsible for pH‐dependent binding of immunoglobulin G (IgG) to neonatal Fc receptor (FcRn) and the results support the current understanding of the salvage mechanism of the antibody by FcRn based on pH‐selective binding. The method can be used to select mutations that change the pH‐dependent binding profiles of proteins and guide the time consuming and expensive protein engineering experiments. As an application of this method, we propose a computational strategy to search for mutations that can alter the pH‐dependent binding behavior of IgG to FcRn with the aim of improving the half‐life of therapeutic antibodies in the target organism. © Proteins 2013. © 2012 Wiley Periodicals, Inc.