| 1. |
- Andersson, David C., 1978-, et al.
(författare)
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Mapping of ligand-binding cavities in proteins
- 2010
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Ingår i: Proteins. - : John Wiley & Sons, Inc. - 0887-3585 .- 1097-0134. ; 78:6, s. 1408-1422
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Tidskriftsartikel (refereegranskat)abstract
- The complex interactions between proteins and small organic molecules (ligands) are intensively studied because they play key roles in biological processes and drug activities. Here, we present a novel approach to characterize and map the ligand-binding cavities of proteins without direct geometric comparison of structures, based on Principal Component Analysis of cavity properties (related mainly to size, polarity, and charge). This approach can provide valuable information on the similarities and dissimilarities, of binding cavities due to mutations, between-species differences and flexibility upon ligand-binding. The presented results show that information on ligand-binding cavity variations can complement information on protein similarity obtained from sequence comparisons. The predictive aspect of the method is exemplified by successful predictions of serine proteases that were not included in the model construction. The presented strategy to compare ligand-binding cavities of related and unrelated proteins has many potential applications within protein and medicinal chemistry, for example in the characterization and mapping of "orphan structures", selection of protein structures for docking studies in structure-based design, and identification of proteins for selectivity screens in drug design programs.
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| 2. |
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| 3. |
- Di Matteo, Adele, et al.
(författare)
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Structural and functional characterization of CcmG from Pseudomonas aeruginosa, a key component of the bacterial cytochrome c maturation apparatus
- 2010
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 78:10, s. 2213-2221
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Tidskriftsartikel (refereegranskat)abstract
- The cytochrome c maturation process is carried out in the bacterial periplasm, where some specialized thiol-disulfide oxidoreductases work in close synergy for the correct reduction of oxidized apocytochrome before covalent heme attachment. We present a structural and functional characterization of the soluble periplasmic domain of CcmG from the opportunistic pathogen P. aeruginosa (Pa-CcmG), a component of the protein machinery involved in cyt c maturation in gram-negative bacteria. X-ray crystallography reveals that Pa-CcmG is a TRX-like protein; high-resolution crystal structures show that the oxidized and the reduced forms of the enzyme are identical except for the active-site disulfide. The standard redox potential was calculated to be E-0' = -0.213 V at pH 7.0; the pK(a) of the active site thiols were pK(a) = 6.13 +/- 0.05 for the N-terminal Cys74 and pK(a) = 10.5 +/- 0.17 for the C-terminal Cys77. Experiments were carried out to characterize and isolate the mixed disulfide complex between Pa-CcmG and Pa-CcmH (the other redox active component of System I in P. aeruginosa). Our data indicate that the target disulfide of this TRX-like protein is not the intramolecular disulfide of oxidized Pa-CcmH, but the intermolecular disulfide formed between Cys28 of Pa-CcmH and DTNB used for the in vitro experiments. This observation suggests that, in vivo, the physiological substrate of Pa-CcmG may be the mixed-disulfide complex between Pa-CcmH and apo-cyt. Proteins 2010; 78:2213-2221. (C) 2010 Wiley-Liss, Inc.
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| 4. |
- Dourado, Daniel F. A. R., et al.
(författare)
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A multiscale approach to predicting affinity changes in protein-protein interfaces
- 2014
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 82:10, s. 2681-2690
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Tidskriftsartikel (refereegranskat)abstract
- Substitution mutations in protein-protein interfaces can have a substantial effect on binding, which has consequences in basic and applied biomedical research. Experimental expression, purification, and affinity determination of protein complexes is an expensive and time-consuming means of evaluating the effect of mutations, making a fast and accurate in silico method highly desirable. When the structure of the wild-type complex is known, it is possible to economically evaluate the effect of point mutations with knowledge based potentials, which do not model backbone flexibility, but these have been validated only for single mutants. Substitution mutations tend to induce local conformational rearrangements only. Accordingly, ZEMu (Zone Equilibration of Mutants) flexibilizes only a small region around the site of mutation, then computes its dynamics under a physics-based force field. We validate with 1254 experimental mutants (with 1-15 simultaneous substitutions) in a wide variety of different protein environments (65 protein complexes), and obtain a significant improvement in the accuracy of predicted Delta Delta G.
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| 5. |
- Friedman, Ran, et al.
(författare)
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On the orientation of the catalytic dyad in aspartic proteases
- 2010
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 78:6, s. 1575-1582
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Tidskriftsartikel (refereegranskat)abstract
- The recent re-refinement of the X-ray structure of apo plasmepsin II from Plasmodium falciparum suggests that the two carboxylate groups in the catalytic dyad are noncoplanar, (Robbins et al., Acta Crystallogr D Biol Crystallogr 2009;65: 294–296) in remarkable contrast with the vast majority of structures of aspartic proteases. Here, evidence for the noncoplanarity of the catalytic aspartates is provided by analysis of multiple explicit water molecular dynamics (MD) simulations of plasmepsin II, human β-secretase, and HIV-protease. In the MD runs of plasmepsin II, the angle between the planes of the two carboxylates of the catalytic dyad is almost always in the range 60°–120°, in agreement with the perpendicular orientation in the re-refined X-ray structure. The noncoplanar arrangement is prevalent also in the β-secretase simulations, as well as in the runs with the inhibitor-bound proteases. Quantum-mechanics calculations provide further evidence that before catalysis the noncoplanar arrangement is favored energetically in eukaryotic aspartic proteases. Remarkably, the coplanar orientation of the catalytic dyad is observed in MD simulations of HIV-protease at 100 K but not at 300 K, which indicates that the noncoplanar arrangement is favored by conformational entropy. This finding suggests that the coplanar orientation in the crystal structures of apo aspartic proteases is promoted by the very low temperature used for data collection (usually around 100 K).
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| 6. |
- Friedman, Ran, et al.
(författare)
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Wild type and mutants of the HET-s(218-289) prion show different flexibility at fibrillar ends : A simulation study
- 2014
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 82:3, s. 399-404
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Tidskriftsartikel (refereegranskat)abstract
- The C-terminal segment (residues 218–289) of the HET-s protein of the filamentous fungus Podosporina anserina is a prion-forming domain. The structural model of the HET-s(218–289) amyloid fibril based on solid-state nuclear magnetic resonance (NMR) restraints shows a β solenoid topology which is comprised of a β-sheet core and interconnecting loops. For the single-point mutants Phe286Ala and Trp287Ala, slower aggregation rates in vitro and loss of prionic infectivity have been reported recently. Here we have used molecular dynamics to compare the flexibility of the mutants and wild type. The simulations, initiated from a trimeric aggregate extracted from the NMR structural model, show structural stability on a 100-ns time scale for wild type and mutants. Analysis of the fluctuations along the simulations reveals that the mutants are less flexible than the wild type in the C-terminal segment at only one of the two external monomers. Analysis of interaction energy and buried accessible surface indicates that residue Phe286 in particular is stabilized in the Trp287Ala mutant. The simulation results provide an atomistic explanation of the suggestion (based on indirect experimental evidence) that flexibility at the protofibril end(s) is required for fibril elongation. Moreover, they provide further evidence that the growth of the HET-s amyloid fibril is directional.
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| 7. |
- Griese, Julia J., et al.
(författare)
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Structure and DNA-binding activity of the Pyrococcus furiosus SMC protein hinge domain
- 2011
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 79:2, s. 558-568
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Tidskriftsartikel (refereegranskat)abstract
- Structural Maintenance of Chromosomes (SMC) proteins are essential for a wide range of processes including chromosome structure and dynamics, gene regulation, and DNA repair. While bacteria and archaea have one SMC protein that forms a homodimer, eukaryotes possess three distinct SMC complexes, consisting of heterodimeric pairs of six different SMC proteins. SMC holocomplexes additionally contain several specific regulatory subunits. The bacterial SMC complex is required for chromosome condensation and segregation. In eukaryotes, this function is carried out by the condensin (SMC2-SMC4) complex. SMC proteins consist of N-terminal and C-terminal domains that fold back onto each other to create an ATPase "head" domain, connected to a central "hinge" domain via a long coiled-coil region. The hinge domain mediates dimerization of SMC proteins and binds DNA. This activity implicates a direct involvement of the hinge domain in the action of SMC proteins on DNA. We studied the SMC hinge domain from the thermophilic archaeon Pyrococcus furiosus. Its crystal structure shows that the SMC hinge domain fold is largely conserved between archaea and bacteria as well as eukarya. Like the eukaryotic condensin hinge domain, the P. furiosus SMC hinge domain preferentially binds single-stranded DNA (ssDNA), but its affinity for DNA is weaker than that of its eukaryotic counterpart, and point mutations reveal that its DNA-binding surface is more confined. The ssDNA-binding activity of its hinge domain might play a role in the DNA-loading process of the prokaryotic SMC complex during replication.
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| 8. |
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| 9. |
- Illergård, Kristoffer, et al.
(författare)
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Why are polar residues within the membrane core evolutionary conserved?
- 2011
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 79:1, s. 79-91
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Tidskriftsartikel (refereegranskat)abstract
- Here, we present a study of polar residues within the membrane core of alpha-helical membrane proteins. As expected, polar residues are less frequent in the membrane than expected. Further, most of these residues are buried within the interior of the protein and are only rarely exposed to lipids. However, the polar groups often border internal water filled cavities, even if the rest of the sidechain is buried. A survey of their functional roles in known structures showed that the polar residues are often directly involved in binding of small compounds, especially in channels and transporters, but other functions including proton transfer, catalysis, and selectivity have also been attributed to these proteins. Among the polar residues histidines often interact with prosthetic groups in photosynthetic-and oxidoreductase-related proteins, whereas pro-lines often are required for conformational changes of the proteins. Indeed, the polar residues in the membrane core are more conserved than other residues in the core, as well as more conserved than polar residues outside the membrane. The reason is twofold; they are often (i) buried in the interior of the protein and (ii) directly involved in the function of the proteins. Finally, a method to identify which polar residues are present within the membrane core directly from protein sequences was developed. Applying the method to the set of all human membrane proteins the prediction indicates that polar residues were most frequent among active transporter proteins and GPCRs, whereas infrequent in families with few transmembrane regions, such as non-GPCR receptors. Proteins 2011; 79: 79-91.
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| 10. |
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| 11. |
- Luo, Jinghui, et al.
(författare)
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Examining the promiscuous phosphatase activity of Pseudomonas aeruginosa arylsulfatase : A comparison to analogous phosphatases
- 2012
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Ingår i: Proteins: Structure, Function and Bioinformatics. - : Wiley. - 1097-0134 .- 0887-3585. ; 80:4, s. 1211-1226
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Tidskriftsartikel (refereegranskat)abstract
- Pseudomonas aeruginosa arylsulfatase (PAS) is a bacterial sulfatase capable ofhydrolyzing a range of sulfate esters. Recently, it has been demonstrated to also show very high proficiency for phosphate ester hydrolysis. Such proficient catalytic promiscuity is significant, as promiscuity has been suggested to play an important role in enzyme evolution. Additionally, a comparative study of the hydrolyses of the p-nitrophenyl phosphate and sulfate monoesters in aqueous solution has demonstrated that despite superficial similarities, the two reactions proceed through markedly different transition states with very different solvation effects, indicating that the requirements for the efficient catalysis of the two reactions by an enzyme will also be very different (and yet they are both catalyzed by thesame active site). This work explores the promiscuous phosphomonoesterase activity ofPAS. Specifically, we have investigated the identity of the most likely base for the initial activation of the unusual formylglycine hydrate nucleophile (which is common to many sulfatases), and demonstrate that a concerted substrate-as-base mechanism is fully consistent with the experimentally observed data. This is very similar to other related systems, and suggests that, as far as the phosphomonoesterase activity of PAS is concerned, the sulfatase behaves like a classical phosphatase, despite the fact that such a mechanism is unlikely to be available to the native substrate (based on pKa considerations and studies of model systems). Understanding such catalytic versatility can be used to design novel artificial enzymes that are far more proficient than the current generation ofdesigner enzymes.
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| 12. |
- Mitternacht, Simon, et al.
(författare)
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Comparing the folding free-energy landscapes of Abeta42 variants with different aggregation properties.
- 2010
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 78:12, s. 2600-2608
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Tidskriftsartikel (refereegranskat)abstract
- The properties of the amyloid-beta peptide that lead to aggregation associated with Alzheimer's disease are not fully understood. This study aims at identifying conformational differences among four variants of full-length Abeta42 that are known to display very different aggregation properties. By extensive all-atom Monte Carlo simulations, we find that a variety of beta-sheet structures with distinct turns are readily accessible for full-length Abeta42. In the simulations, wild type (WT) Abeta42 preferentially populates two major classes of conformations, either extended with high beta-sheet content or more compact with lower beta-sheet content. The three mutations studied alter the balance between these classes. Strong mutational effects are observed in a region centered at residues 23-26, where WT Abeta42 tends to form a turn. The aggregation-accelerating E22G mutation associated with early onset of Alzheimer's disease makes this turn region conformationally more diverse, whereas the aggregation-decelerating F20E mutation has the reverse effect, and the E22G/I31E mutation reduces the turn population. Comparing results for the four Abeta42 variants, we identify specific conformational properties of residues 23-26 that might play a key role in aggregation. Proteins 2010. (c) 2010 Wiley-Liss, Inc.
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| 13. |
- Moretti, Rocco, et al.
(författare)
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Community-wide evaluation of methods for predicting the effect of mutations on protein-protein interactions
- 2013
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 81:11, s. 1980-1987
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Tidskriftsartikel (refereegranskat)abstract
- Community-wide blind prediction experiments such as CAPRI and CASP provide an objective measure of the current state of predictive methodology. Here we describe a community-wide assessment of methods to predict the effects of mutations on protein-protein interactions. Twenty-two groups predicted the effects of comprehensive saturation mutagenesis for two designed influenza hemagglutinin binders and the results were compared with experimental yeast display enrichment data obtained using deep sequencing. The most successful methods explicitly considered the effects of mutation on monomer stability in addition to binding affinity, carried out explicit side-chain sampling and backbone relaxation, evaluated packing, electrostatic, and solvation effects, and correctly identified around a third of the beneficial mutations. Much room for improvement remains for even the best techniques, and large-scale fitness landscapes should continue to provide an excellent test bed for continued evaluation of both existing and new prediction methodologies. Proteins 2013; 81:1980-1987.
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| 14. |
- Nivón, Lucas G, et al.
(författare)
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Automating human intuition for protein design
- 2014
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 82:5, s. 858-866
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Tidskriftsartikel (refereegranskat)abstract
- In the design of new enzymes and binding proteins, human intuition is often used to modify computationally designed amino acid sequences prior to experimental characterization. The manual sequence changes involve both reversions of amino acid mutations back to the identity present in the parent scaffold and the introduction of residues making additional interactions with the binding partner or backing up first shell interactions. Automation of this manual sequence refinement process would allow more systematic evaluation and considerably reduce the amount of human designer effort involved. Here we introduce a benchmark for evaluating the ability of automated methods to recapitulate the sequence changes made to computer-generated models by human designers, and use it to assess alternative computational methods. We find the best performance for a greedy one-position-at-a-time optimization protocol that utilizes metrics (such as shape complementarity) and local refinement methods too computationally expensive for global Monte Carlo (MC) sequence optimization. This protocol should be broadly useful for improving the stability and function of designed binding proteins.
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| 15. |
- Peters, Christoph, et al.
(författare)
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Why is the biological hydrophobicity scale more accurate than earlier experimental hydrophobicity scales?
- 2014
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 82:9, s. 2190-2198
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Tidskriftsartikel (refereegranskat)abstract
- The recognition of transmembrane helices by the translocon is primarily guided by the average hydrophobicity of the protential transmembrane helix, However, he exact hydrophobicity of each amino acid can he identified in several diferent ways. The free energy of transfer for amino acid analogues between a hydrophobic media, for example, octanol and water can be measured or obtained from simulations, the hydrophobicity can also be estimated by statistical properties from known transmembrane segments and finally the contribution of each amino acid type for the probability of traitslocon recognition has recently been measured directly. Although these scales correlate quite well, there are dear differences between them and it is not well understood which scale represents neither the biology best nor what the differences are. Here, we try to provide some answers to this by studying the ability of different scales to recognize transmembrane helices and predict the topology of transmembrane proteins. From this analysis it is clear that the biological hydrophobicity scale as well scales created from statistical analysis of membrane helices perform better than earlier experimental scales that are mainly based on measurements of amino acid analogs and not directly on transmeiribrane helix recognition. Using these results we identified the properties of the scales that perform better than other scales. We find, for instance, that the better performing scales consider proline more hydrophilic. This shows that tratismembrarie recognition is not only governed by pure hydrophobicity but also by the helix preferences for amino acids, as praline is a strong helix breaker.
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| 16. |
- Repic, Matej, et al.
(författare)
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Empirical valence bond simulations of the hydride transfer step in the monoamine oxidase B catalyzed metabolism of dopamine
- 2014
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 82:12, s. 3347-3355
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Tidskriftsartikel (refereegranskat)abstract
- Monoamine oxidases (MAOs) A and B are flavoenzymes responsible for the metabolism of biogenic amines such as dopamine, serotonin and noradrenaline. In this work, we present a comprehensive study of the rate-limiting step of dopamine degradation by MAO B, which consists in the hydride transfer from the methylene group of the substrate to the flavin moiety of the FAD prosthetic group. This article builds on our previous quantum chemical study of the same reaction using a cluster model (Vianello et al., Eur J Org Chem 2012; 7057), but now considering the full dimensionality of the hydrated enzyme with extensive configurational sampling. We show that MAO B is specifically tuned to catalyze the hydride transfer step from the substrate to the flavin moiety of the FAD prosthetic group and that it lowers the activation barrier by 12.3 kcal mol(-1) compared to the same reaction in aqueous solution, a rate enhancement of more than nine orders of magnitude. Taking into account the deprotonation of the substrate prior to the hydride transfer reaction, the activation barrier in the enzyme is calculated to be 16.1 kcal mol(-1), in excellent agreement with the experimental value of 16.5 kcal mol(-1). Additionally, we demonstrate that the protonation state of the active site residue Lys296 does not have an influence on the hydride transfer reaction.
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| 17. |
- Todde, Guido, et al.
(författare)
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Glucose oxidase from Penicillium amagasakiense : Characterization of the transition state of its denaturation from molecular dynamics simulations
- 2014
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 82:10, s. 2353-2363
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Tidskriftsartikel (refereegranskat)abstract
- Glucose oxidase (GOx) is a flavoenzyme having applications in food and medical industries. However, GOx, as many other enzymes when extracted from the cells, has relatively short operational lifetimes. Several recent studies (both experimental and theoretical), carried out on small proteins (or small fractions of large proteins), show that a detailed knowledge of how the breakdown process starts and proceeds on molecular level could be of significant help to artificially improve the stability of fragile proteins. We have performed extended molecular dynamics (MD) simulations to study the denaturation of GOx (a protein dimer containing nearly 1200 amino acids) to identify weak points in its structure and in this way gather information to later make it more stable, for example, by mutations. A denaturation of a protein can be simulated by increasing the temperature far above physiological temperature. We have performed a series of MD simulations at different temperatures (300, 400, 500, and 600 K). The exit from the protein's native state has been successfully identified with the clustering method and supported by other methods used to analyze the simulation data. A common set of amino acids is regularly found to initiate the denaturation, suggesting a moiety where the enzyme could be strengthened by a suitable amino acid based modification.
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| 18. |
- Xue, Bin, et al.
(författare)
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Abundance and functional roles of intrinsic disorder in allergenic proteins and allergen representative peptides
- 2011
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Ingår i: Proteins. - : Wiley. - 0887-3585 .- 1097-0134. ; 79:9, s. 2595-2606
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Tidskriftsartikel (refereegranskat)abstract
- The pathological process of allergies generally involves an initial activation of certain immune cells, tied to an ensuing inflammatory reaction on renewed contact with the allergen. In IgE-mediated hypersensitivity, this typically occurs in response to otherwise harmless food-or air-borne proteins. As some members of certain protein families carry special properties that make them allergenic, exploring protein allergens at the molecular level is instrumental to an improved understanding of the disease mechanisms, including the identification of relevant antigen features. For this purpose, we inspected a previously identified set of allergen representative peptides (ARPs) to scrutinize protein intrinsic disorder. The resulting study presented here focused on the association between these ARPs and protein intrinsic disorder. In addition, the connection between the disorder-enriched ARPs and UniProt functional keywords was considered. Our analysis revealed that similar to 20% of the allergen peptides are highly disordered, and that similar to 77% of ARPs are either located within disordered regions of corresponding allergenic proteins or show more disorder/flexibility than their neighbor regions. Furthermore, among the subset of allergenic proteins, similar to 70% of the predicted molecular recognition features (MoRFs that consist of short interactive disordered regions undergoing disorder-to-order transitions at interaction with binding partners) were identified as ARPs. These results suggest that intrinsic disorder and MoRFs may play functional roles in IgE-mediated allergy.
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| 19. |
- Genheden, Samuel, et al.
(författare)
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Comparison of end-point continuum-solvation methods for the calculation of protein-ligand binding free energies.
- 2012
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Ingår i: Proteins. - : Wiley. - 0887-3585. ; 80:5, s. 1326-1342
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Tidskriftsartikel (refereegranskat)abstract
- We have compared the predictions of ligand-binding affinities from several methods based on end-point molecular dynamics simulations and continuum solvation, i.e. methods related to MM/PBSA (molecular mechanics combined with Poisson-Boltzmann and surface area solvation). Two continuum-solvation models were considered, viz. the Poisson-Boltzmann (PB) and generalised Born (GB) approaches. The non-electrostatic energies were also obtained in two different ways, viz. either from the sum of the bonded, van der Waals, non-polar solvation energies, and entropy terms (as in MM/PBSA), or from the scaled protein-ligand van der Waals interaction energy (as in the linear interaction energy approach, LIE). Three different approaches to calculate electrostatic energies were tested, viz. the sum of electrostatic interaction energies and polar solvation energies, obtained either from a single simulation of the complex or from three independent simulations of the complex, the free protein, and the free ligand, or the linear-response approximation (LRA). Moreover, we investigated the effect of scaling the electrostatic interactions by an effective internal dielectric constant of the protein (ε(int) ). All these methods were tested on the binding of seven biotin analogues to avidin and nine 3-amidinobenzyl-1H-indole-2-carboxamide inhibitors to factor Xa. For avidin, the best results were obtained with a combination of the LIE non-electrostatic energies with the MM+GB electrostatic energies from a single simulation, using ε(int) = 4. For fXa, standard MM/GBSA, based on one simulation and using ε(int) = 4-10 gave the best result. The optimum internal dielectric constant seems to be slightly higher with PB than with GB solvation. Proteins 2012. © 2012 Wiley-Liss, Inc.
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| 20. |
- Jonsson, Sigurdur, et al.
(författare)
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Distinct phases of free α-synuclein - A Monte Carlo study.
- 2012
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Ingår i: Proteins. - : Wiley. - 0887-3585. ; 80:9, s. 2169-2177
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Tidskriftsartikel (refereegranskat)abstract
- The α-synuclein protein (αS), implicated in Parkinson's disease (PD), shows conformational versatility. It aggregates into β-sheet-rich fibrils, occurs in helical membrane-bound forms, is disordered as a free monomer, and has recently been suggested to have a folded helical tetramer as its main physiological form. Here we use implicit solvent all-atom Monte Carlo (MC) methods to explore the conformational ensemble sampled by the free αS monomer. We analyze secondary-structure propensities, size and topological properties, and compare with existing experimental data. Our study suggests that free αS has two distinct phases. One phase has the expected disordered character. The other phase also shows large conformational variability. However, in this phase, the β-strand content is substantial, and the backbone fold shows statistical similarities with that in αS fibrils. Presence of this phase is consistent with data from low-temperature experiments. Conversion of disordered αS to this fibril-like form requires the crossing of a rather large apparent free-energy barrier. Proteins 2012. © 2012 Wiley Periodicals, Inc.
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| 21. |
- Kurut Sabanoglu, Anil, et al.
(författare)
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Role of histidine for charge regulation of unstructured peptides at interfaces and in bulk.
- 2014
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Ingår i: Proteins. - : Wiley. - 0887-3585. ; 82:4, s. 657-667
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Tidskriftsartikel (refereegranskat)abstract
- Histidine rich, unstructured peptides adsorb to charged interfaces such as mineral surfaces and microbial cell membranes. At a molecular level, we investigate the adsorption mechanism as a function of pH, salt, and multivalent ions showing that (1) proton charge fluctuations are - in contrast to the majority of proteins - optimal at neutral pH, promoting electrostatic interactions with anionic surfaces through charge regulation, and (2) specific zinc(II)-histidine binding competes with protons and ensures an unusually constant charge distribution over a broad pH interval. In turn this further enhances surface adsorption. Our analysis is based on atomistic molecular dynamics simulations, coarse grained Metropolis Monte Carlo, and classical polymer density functional theory. This multi-scale modelling provides a consistent picture in good agreement with experimental data on Histatin 5, an antimicrobial salivary peptide. Biological function is discussed and we suggest that charge regulation is a significant driving force for the remarkably robust activity of histidine rich antimicrobial peptides.
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| 22. |
- Rath, Emma M, et al.
(författare)
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Small-angle X-ray scattering of BAMLET at pH 12 : a complex of α-lactalbumin and oleic acid
- 2014
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Ingår i: Proteins. - : Wiley. - 0887-3585. ; 82:7, s. 8-1400
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Tidskriftsartikel (refereegranskat)abstract
- BAMLET (Bovine Alpha-lactalbumin Made LEthal to Tumors) is a member of the family of the HAMLET-like complexes, a novel class of protein-based anti-cancer complexes that incorporate oleic acid and deliver it to cancer cells. Small angle X-ray scattering (SAXS) was performed on the complex at pH 12, examining the high pH structure as a function of oleic acid added. The SAXS data for BAMLET species prepared with a range of oleic acid concentrations indicate extended, irregular, partially unfolded protein conformations that vary with the oleic acid concentration. Increases in oleic acid concentration correlate with increasing radius of gyration without an increase in maximum particle dimension, indicating decreasing protein density. The models for the highest oleic acid content BAMLET indicate an unusual coiled elongated structure that contrasts with apo-α-lactalbumin at pH 12, which is an elongated globular molecule, suggesting that oleic acid inhibits the folding or collapse of the protein component of BAMLET to the globular form. Circular dichroism of BAMLET and apo-α-lactalbumin was performed and the results suggest that α-lactalbumin and BAMLET unfold in a continuum of increasing degree of unfolded states. Taken together, these results support a model in which BAMLET retains oleic acid by non-specific association in the core of partially unfolded protein, and represent a new type of lipoprotein structure.
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| 23. |
- von Schantz, Laura, et al.
(författare)
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Polar interactions with branching xyloses and CH-π interactions define carbohydrate binding module recognition of xyloglucan
- 2014
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Ingår i: Proteins. - : Wiley. - 0887-3585. ; 82:12, s. 3466-3475
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Tidskriftsartikel (refereegranskat)abstract
- Engineering of novel carbohydrate-binding proteins that can be utilized in various biochemical and biotechnical applications would benefit from a deeper understanding of the biochemical interactions that determine protein-carbohydrate specificity. In an effort to understand further the basis for specificity we present the crystal structure of the multi-specific carbohydrate-binding module (CBM) X-2 L110F bound to a branched oligomer of xyloglucan (XXXG). X-2 L110F is an engineered CBM that can recognize xyloglucan, xylans and β-glucans. The structural observations of the present study compared with previously reported structures of X-2 L110F in complex with linear oligomers, show that the π-surface of a phenylalanine, F110, allows for interactions with hydrogen atoms on both linear (xylopentaose and cellopentaose) and branched ligands (XXXG). Furthermore, X-2 L110F is shown to have a relatively flexible binding cleft, as illustrated in binding to XXXG. This branched ligand requires a set of reorientations of protein side chains Q72, N31, and R142, although these residues have previously been determined as important for binding to xylose oligomers by mediating polar contacts. The loss of these polar contacts is compensated for in binding to XXXG by polar interactions mediated by other protein residues, T74, R115, and Y149, which interact mainly with the branching xyloses of the xyloglucan oligomer. Taken together, the present study illustrates in structural detail how CH-π interactions can influence binding specificity and that flexibility is a key feature for the multi-specificity displayed by X-2 L110F, allowing for the accommodation of branched ligands.
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| 24. |
- Xie, Yongjing, et al.
(författare)
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Electrostatic interactions play an essential role in the binding of oleic acid with a-lactalbumin in the HAMLET-like complex: A study using charge-specific chemical modifications
- 2013
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Ingår i: Proteins. - : Wiley. - 0887-3585. ; 81:1, s. 1-17
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Tidskriftsartikel (refereegranskat)abstract
- Human a-lactalbumin made lethal to tumor cells (HAMLET) and its analogs are partially unfolded protein-oleic acid (OA) complexes that exhibit selective tumoricidal activity normally absent in the native protein itself. To understand the nature of the interaction between protein and OA moieties, charge-specific chemical modifications of lysine side chains involving citraconylation, acetylation, and guanidination were employed and the biophysical and biological properties were probed. Upon converting the original positively-charged lysine residues to negatively-charged citraconyl or neutral acetyl groups, the binding of OA to protein was eliminated, as were any cytotoxic activities towards osteosarcoma cells. Retention of the positive charges by converting lysine residues to homoarginine groups (guanidination); however, yielded unchanged binding of OA to protein and identical tumoricidal activity to that displayed by the wild-type a-lactalbumin-oleic acid complex. With the addition of OA, the wild-type and guanidinated a-lactalbumin proteins underwent substantial conformational changes, such as partial unfolding, loss of tertiary structure, but retention of secondary structure. In contrast, no significant conformational changes were observed in the citraconylated and acetylated a-lactalbumins, most likely because of the absence of OA binding. These results suggest that electrostatic interactions between the positively-charged basic groups on a-lactalbumin and the negatively-charged carboxylate groups on OA molecules play an essential role in the binding of OA to a-lactalbumin and that these interactions appear to be as important as hydrophobic interactions. Proteins 2013. (c) 2012 Wiley Periodicals, Inc.
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