bluuebunny/crossref_metadata_2025_split huggingface dataset:
Protein–protein binding is often associated with changes in protonation state
Authors: Mason Aaron C. , Jensen Jan H. 
Published: October 2007

AbstractpKa values of ionizable residues have been calculated using the PROPKA method and structures of 75 protein–protein complexes and their corresponding free forms. These pKa values were used to compute changes in protonation state of individual residues, net changes in protonation state of the complex relative to the uncomplexed proteins, and the correction to a binding energy calculated assuming standard protonation states at pH 7. For each complex, two different structures for the uncomplexed form of the proteins were used: the X‐ray structures determined for the proteins in the absence of the other protein and the individual protein structures taken from the structure of the complex (referred to as unbound and bound structures, respectively). In 28 and 77% of the cases considered here, protein–protein binding is accompanied by a complete (>95%) or significant (>50%) change in protonation state of at least one residue using unbound structures. Furthermore, in 36 and 61% of the cases, protein–protein binding is accompanied by a complete or significant net change in protonation state of the complex relative to the separated monomers. Using bound structures, the corresponding values are 12, 51, 20, and 48%. Comparison to experimental data suggest that using unbound and bound structures lead to over‐ and underestimation of binding‐induced protonation state changes, respectively. Thus, we conclude that protein–protein binding is often associated with changes in protonation state of amino acid residues and with changes in the net protonation state of the proteins. The pH‐dependent correction to the binding energy contributes at least one order of magnitude to the binding constant in 45 and 23%, using unbound and bound structures, respectively. Proteins 2008. © 2007 Wiley‐Liss, Inc.
Osmolyte effect on the stability and folding of a hyperthermophilic protein
Authors: Mukaiyama Atsushi , Koga Yuichi , Takano Kazufumi , Kanaya Shigenori 
Published: October 2007

AbstractProteins are known to be stabilized by naturally occurring osmolytes such as amino acids, sugars, and methylamines. Here, we examine the effect of trimethylamine‐N‐oxide (TMAO) on the conformational stability of ribonuclease HII from a hyperthermophile, Thermococcus kodakaraensis (Tk‐RNase HII), which inherently possesses high conformational stability. Heat‐ and guanidine hydrochloride‐induced unfolding experiments demonstrated that the conformational stability of Tk‐RNase HII in the presence of 0.5M TMAO was higher than that in the absence of TMAO at all examined temperatures. TMAO affected the unfolding and refolding kinetics of Tk‐RNase HII to a similar extent. These results indicate that proteins are universally stabilized by osmolytes, regardless of their robustness, and suggest a stabilization mechanism by osmolytes, caused by the unfavorable interaction of osmolytes with protein backbones in the denatured state. Our results also imply that the basic protein folding principle is not dependent on protein stability and evolution. Proteins 2008. © 2007 Wiley‐Liss, Inc.
Large‐scale characteristics of the energy landscape in protein–protein interactions
Authors: O'Toole Nicholas , Vakser Ilya A. 
Published: October 2007

AbstractCharacterization of intermolecular energy landscapes in protein–protein interactions is important for understanding the mechanisms of these interactions as well as for designing better protein docking methods. A simplified representation of the landscape was used for a systematic study of its large‐scale characteristics in a large nonredundant dataset of protein complexes. The focus of the study is on the basic features of the low‐resolution energy basins and their distribution on the landscape. The results clearly show that, in general, the number of such basins is small, these basins are well formed, correlated with actual binding modes, and the pattern of basins distribution depends on the type of the complex. For docking studies, the results suggest that adequate starting points for the structural refinement are detectable by low‐resolution procedures and the number of such starting points is relatively small. Proteins 2008. © 2007 Wiley‐Liss, Inc.
Conformational selection in silico: Loop latching motions and ligand binding in enzymes
Authors: Wong Sergio , Jacobson Matthew P. 
Published: October 2007

AbstractLigand binding frequently induces significant conformational changes in a protein receptor. Understanding and predicting such conformational changes represent an important challenge for computational biology, including applications to structure‐based drug design. We describe an approach to this problem based on the assumption that the holo state is at least transiently populated in the absence of a ligand; this hypothesis has been referred to as “conformational selection.” Here, we apply a method that tests this hypothesis on a challenging class of ligand‐induced conformational changes, which we refer to as loop latching: the closing of a loop around an active site that sequesters the ligand from solvent. The method uses a combination of replica exchange molecular dynamics and a loop prediction algorithm to generate low‐energy loop structures, and docking to select the conformation appropriate for binding a particular ligand. On a test set of six proteins, it yields loop structures including hololike conformations, generally below 2 Å RMSD from the liganded structure, for loops that span up to 15 residues. Docking serves as a stringent test of the predictions. In five of the six cases, the predicted loop conformations improve the ranks of cognate ligands relative to using the apo structure, although the results remain, in most cases, significantly worse than using a holo structure. The poses of the cognate ligands are correct in four of the six test cases, while they are correct for five of the six using a holo structure. Proteins 2008. © 2007 Wiley‐Liss, Inc.
Engineering proteins with tunable thermodynamic and kinetic stabilities
Authors: Pey Angel L. , Rodriguez‐Larrea David , Bomke Susanne , Dammers Susanne , Godoy‐Ruiz Raquel , Garcia‐Mira Maria M. , Sanchez‐Ruiz Jose M. 
Published: October 2007

AbstractIt is widely recognized that enhancement of protein stability is an important biotechnological goal. However, some applications at least, could actually benefit from stability being strongly dependent on a suitable environment variable, in such a way that enhanced stability or decreased stability could be realized as required. In therapeutic applications, for instance, a long shelf‐life under storage conditions may be convenient, but a sufficiently fast degradation of the protein after it has performed the planned molecular task in vivo may avoid side effects and toxicity. Undesirable effects associated to high stability are also likely to occur in food‐industry applications. Clearly, one fundamental factor involved here is the kinetic stability of the protein, which relates to the time‐scale of the irreversible denaturation processes and which is determined to some significant extent by the free‐energy barrier for unfolding (the barrier that “separates” the native state from the highly‐susceptible‐to‐irreversible‐alterations nonnative states). With an appropriate experimental model, we show that strong environment‐dependencies of the thermodynamic and kinetic stabilities can be achieved using robust protein engineering. We use sequence‐alignment analysis and simple computational electrostatics to design stabilizing and destabilizing mutations, the latter introducing interactions between like charges which are screened out at high salt. Our design procedures lead naturally to mutating regions which are mostly unstructured in the transition state for unfolding. As a result, the large salt effect on the thermodynamic stability of our consensus plus charge‐reversal variant translates into dramatic changes in the time‐scale associated to the unfolding barrier: from the order of years at high salt to the order of days at low salt. Certainly, large changes in salt concentration are not expected to occur in biological systems in vivo. Hence, proteins with strong salt‐dependencies of the thermodynamic and kinetic stabilities are more likely to be of use in those cases in which high‐stability is required only under storage conditions. A plausible scenario is that inclusion of high salt in liquid formulations will contribute to a long protein shelf‐life, while the lower salt concentration under the conditions of the application will help prevent the side effects associated with high‐stability which may potentially arise in some therapeutic and food‐industry applications. From a more general viewpoint, this work shows that consensus engineering and electrostatic engineering can be readily combined and clarifies relevant aspects of the relation between thermodynamic stability and kinetic stability in proteins. Proteins 2008. © 2007 Wiley‐Liss, Inc.
Prediction of RNA binding sites in a protein using SVM and PSSM profile
Authors: Kumar Manish , Gromiha M. Michael , Raghava G. P. S. 
Published: October 2007

AbstractRNA‐binding proteins (RBPs) play key roles in post‐transcriptional control of gene expression, which, along with transcriptional regulation, is a major way to regulate patterns of gene expression during development. Thus, the identification and prediction of RNA binding sites is an important step in comprehensive understanding of how RBPs control organism development. Combining evolutionary information and support vector machine (SVM), we have developed an improved method for predicting RNA binding sites or RNA interacting residues in a protein sequence. The prediction models developed in this study have been trained and tested on 86 RNA binding protein chains and evaluated using fivefold cross validation technique. First, a SVM model was developed that achieved a maximum Matthew's correlation coefficient (MCC) of 0.31. The performance of this SVM model further improved the MCC from 0.31 to 0.45, when multiple sequence alignment in the form of PSSM profiles was used as input to the SVM, which is far better than the maximum MCC achieved by previous methods (0.41) on the same dataset. In addition, SVM models were also developed on an alternative dataset that contained 107 RBP chains. Utilizing PSSM as input information to the SVM, the training/testing on this alternate dataset achieved a maximum MCC of 0.32. Conclusively, the prediction performance of SVM models developed in this study is better than the existing methods on the same datasets. A web server ‘Pprint’ was also developed for predicting RNA binding residues in a protein sequence which is freely available at http://www.imtech.res.in/raghava/pprint/. Proteins 2008. © 2007 Wiley‐Liss, Inc.
The peculiar structural features of kiwi fruit pectin methylesterase: Amino acid sequence, oligosaccharides structure, and modeling of the interaction with its natural proteinaceous inhibitor
Authors: Ciardiello M. Antonietta , D'Avino Rossana , Amoresano Angela , Tuppo Lisa , Carpentieri Andrea , Carratore Vito , Tamburrini Maurizio , Giovane Alfonso , Pucci Piero , Camardella Laura 
Published: October 2007

AbstractPectin methylesterase (PME) from kiwi fruit (Actinidia deliciosa) is a glycoprotein, showing an apparent molecular mass of 50 kDa upon size exclusion chromatography and SDS‐PAGE. The primary structure, elucidated by direct sequencing of the protein, comprises 321 amino acid residues providing a molecular mass of 35 kDa. The protein has an acetylated Thr residue at the amino terminus and five N‐glycosylation consensus sequences, four of which are actually glycosylated. A careful investigation of the oligosaccharide structures demonstrated that PME glycans belong to complex type oligosaccharides essentially consisting of xylosylated polyfucosylated biantennary structures. Alignment with known mature plant PME sequences indicates that the postulated active site residues are conserved. Kiwi PME activity is inhibited following the interaction with the proteinaceous inhibitor PMEI, isolated from the same source. Gel‐filtration experiments show that kiwi PME/PMEI complex is stable in a large pH range and dissociates only at pH 10.0. Modeling of the interaction with the inhibitor was performed by using the crystal structure of the complex between kiwi PMEI and tomato PME as a template. The model shows that the binding site is the same reported for tomato PME. However, additional salt link interactions are found to connect the external loops of kiwi PME to PMEI. This finding may explain the higher pH stability of the complex formed by the two kiwi proteins respect to that formed by PMEI and tomato PME. Proteins 2008. © 2007 Wiley‐Liss, Inc.
Altering the laccase functionality by in vivo assembly of mutant libraries with different mutational spectra
Authors: Zumárraga Miren , Camarero Susana , Shleev Sergey , Martínez‐Arias Arturo , Ballesteros Antonio , Plou Francisco J. , Alcalde Miguel 
Published: October 2007

AbstractThe generation of diversity for directed protein evolution experiments shows an important bottleneck in the in vitro random mutagenesis protocols. Most of them are biased towards specific changes that eventually confer a predicted and conservative mutational spectrum, limiting the exploration of the vast protein space. The current work describes a simple methodology to in vivo recombine mutant libraries with different nucleotide bias created by in vitro methods. This in vivo assembly was based on the accurate physiology of Saccharomyces cerevisiae, which as host, provided its high homologous recombination frequency to shuffle the libraries in a nonmutagenic way. The fungal thermophilic laccase from Myceliophthora thermophila expressed in S. cerevisiae was submitted to this protocol under the selective pressure of high concentrations of organic solvents. Mutant 2E9 with ∼3‐fold better kinetics than parent type showed two consecutive amino acid changes (G614D ‐GGC/GAC‐ and E615K ‐GAG/AAG‐) because of the in vivo shuffling of the mutant libraries. Both mutations are located in the C‐terminal tail that is specifically processed at the Golgi during the maturation of the protein by the Kex2 protease. Notoriously, the oxygen consumption at the T2/T3 trinuclear copper cluster was altered and the catalytic copper at the T1 site was perturbed showing differences in its redox potential and geometry. The change in the isoelectric point of C‐terminal extension upon mutations seems to affect the folding of the protein at the posttranslational processing steps providing new insights in the significance of the C‐terminal tail for the functionality of the ascomycete laccases. Proteins 2008. © 2007 Wiley‐Liss, Inc.
Design and analysis of structure–activity relationship of novel antimicrobial peptides derived from the conserved sequence of cecropin
Authors: Hao Gang , Shi Yong‐Hui , Han Jing‐Hui , Li Qi‐Hui , Tang Ya‐Li , Le Guo‐Wei 
Published: October 2007

AbstractWe have de novo designed four antimicrobial peptides AMP‐A/B/C/D, the 51‐residues peptides, which are based on the conserved sequence of cecropin. In the present study, the four peptides were chemically synthesized and their activities assayed. Their secondary structure, amphipathic property, electric field distribution and transmembrane domain were subsequently predicted by bioinformatics tools. Finally, the structure–activity relationship was analyzed from the results of activity experiments and prediction. The results of activity experiments indicated that AMP‐B/C/D clearly possessed excellent broad‐spectrum activity against bacteria, whereas AMP‐A was almost inactive against most of the bacterial strains tested. AMP‐B/C/D showed more potent activity against Gram‐positive bacteria than against Gram‐negative bacteria. By utilizing bioinformatics analysis tools, we found that the secondary structure of the four cation peptides was mainly α‐helix, and the result of CD spectrum also displayed that all the peptides had considerable α‐helix in the presence of either 50% TFE or SDS micelles. AMP‐C showed much better activity than other peptides against most of the bacteria tested, owing to its remarkable cation property and the amphipathic character of its N‐terminal. The study of structure–activity relationship of the designed peptides confirmed that amphipathic structure and high net positive charge were prerequisites for maintaining their activities. Copyright © 2007 European Peptide Society and John Wiley & Sons, Ltd.
Synthesis and ex vivo profiling of chemically modified cytomegalovirus CMVpp65 epitopes
Authors: Jones Matthew A. , Notta Jatinder K. , Cobbold Mark , Palendira Mainthan , Hislop Andrew D. , Wilkie John , Snaith John S. 
Published: October 2007

AbstractThe effect of substituting unnatural hydrophobic amino acids into the critical MHC binding residues of an HLA‐A*0201‐restricted cytomegalovirus CMVpp65 epitope, NLVPMVATV, has been investigated. A new set of peptides containing the amino acids tert‐butyl glycine (Tgl), cyclohexyl glycine (Chg), neo‐pentyl glycine (Npg), cyclohexyl alanine (Cha) and cyclo leucine (Cyl), at either position 2, to mimic Leu, or position 9, to mimic Val, have been synthesised. Immunological profiling using class I MHC stabilisation assays to assess MHC binding affinity, and enzyme‐linked immunospot (ELISPOT) assays to assess the ability of the modified peptides to re‐stimulate a specific cytotoxic T‐lymphocyte (CTL) response, compared to the native epitope, have been performed. It was found that the majority of the unnatural substitutions resulted in a decrease in either HLA‐A*0201 binding affinity or cytotoxic T‐cell activity. However, the HLA‐A*0201 binding affinity was unrelated to the ability to re‐stimulate a T‐cell response. Minimisation and molecular dynamics studies proved helpful in dissecting the ELISPOT responses. Two principal peptide binding modes were found by minimisation, designated kinked and straight. Peptides that bound in a kinked conformation were poor at re‐stimulating a T‐cell response. Of the peptides that bound in a straight conformation, molecular dynamics (MD) simulations revealed that those capable of re‐stimulating the strongest responses had the greatest degree of flexibility (as determined by RMSD values across the MD simulation) around the P6 residue, one of the residues important for T‐cell receptor recognition. Copyright © 2007 European Peptide Society and John Wiley & Sons, Ltd.
Structure–activity relationships for the cardiotropic action of the Led‐NPF‐I peptide in the beetles Tenebrio molitor and Zophobas atratus
Authors: Marciniak Paweł , Grodecki Sebastian , Konopińska Danuta , Rosiński Grzegorz 
Published: October 2007

AbstractWe have examined the effects of the Led‐NPF‐I peptide (Ala‐Arg‐Gly‐Pro‐Gln‐Leu‐Arg‐Leu‐Arg‐Phe‐amide) and a series of ten analogues on the heart contractile activity of Tenebrio molitor and Zophobas atratus, and the structure–activity relationships for cardioactive action of Led‐NPF‐I were established. A video microscopy technique and computer‐based method of data acquisition and analysis were used to study the action of the peptides on continuously perfused heart preparations. Cardiac activity was progressively inhibited by Led‐NPF‐I when the peptide concentrations were increased from 10−9 to 10−5 M. Substitution of the L‐proline residue at position 4 of the native peptide with hydroxyproline, valine or D‐proline caused a loss of cardioinhibitory activity. Also, replacement of arginine residues at all three positions 2, 7 and 9 with another basic amino acid histidine, reduces cardioinhibitory action of Led‐NPF‐I. Some modifications of the C‐terminal residues, as the Phe(4‐NO2)‐, Phe(4‐NH2)‐ and Phe(4‐NMe2)‐analogues, resulted in agonistic peptides with biological activity similar to that of the native peptide. However, three other C‐terminal analogues tested [Tyr10]‐, [D‐Phe10]‐Led‐NPF‐I, and Ala‐Arg‐Gly‐Pro‐Gln‐Leu‐Arg‐Leu‐Arg‐Phe‐OH were inactive in the heart bioassay, which suggests that this end of the amino acid chain may play an important role in bioactivity and interaction of the native peptide with its receptor on the myocardium. Copyright © 2007 European Peptide Society and John Wiley & Sons, Ltd.
A Mini Review of Mammalian Toxicity (Q)SAR Models
Authors: Tsakovska Ivanka , Lessigiarska Iglika , Netzeva Tatiana , Worth Andrew P. 
Published: October 2007

AbstractThis paper is based on an in‐depth review performed by the European Chemicals Bureau of the European Commission's Joint Research Centre in support of the development of technical guidance for the implementation of the Registration, Evaluation, Authorisation and Restriction of Chemicals (REACH) legislation, and is one of a series of minireviews in this journal. Compared with the QSAR modelling of ecotoxicity, the modelling of mammalian toxicity is more complicated. This is reflected by the relatively small number of QSAR studies on mammalian toxicity published in the literature. The main problem is related to wide variations in the accuracy of the in vivo data, in the organisms used, and also in the complexity and poor understanding of the mechanisms involved, especially for long‐term effects. Nevertheless, the efforts already made to develop QSAR models for mammalian toxicity demonstrate the usefulness of the approach not only for predictive purposes but also for a better understanding of the multiple mechanisms involved in the toxicity. However, there is a need to better characterise existing models in accordance with the OECD QSAR validation principles, to develop new models, and to investigate the applicabilities of individual models in the regulatory context, such as for the purposes of the REACH legislation.
Analysis of longitudinal laboratory data in the presence of common selection mechanisms: A view toward greater emphasis on pre‐marketing pharmaceutical safety
Authors: Schildcrout Jonathan S. , Jenkins Cathy A. , Ostroff Jack H. , Gillen Daniel L. , Harrell Frank E. , Trost Donald C. 
Published: October 2007

AbstractPharmaceutical safety has received substantial attention in the recent past; however, longitudinal clinical laboratory data routinely collected during clinical trials to derive safety profiles are often used ineffectively. For example, these data are frequently summarized by comparing proportions (between treatment arms) of participants who cross pre‐specified threshold values at some time during follow‐up. This research is intended, in part, to encourage more effective utilization of these data by avoiding unnecessary dichotomization of continuous data, acknowledging and making use of the longitudinal follow‐up, and combining data from multiple clinical trials. However, appropriate analyses require careful consideration of a number of challenges (e.g. selection, comparability of study populations, etc.). We discuss estimation strategies based on estimating equations and maximum likelihood for analyses in the presence of three response history‐dependent selection mechanisms: dropout, follow‐up frequency, and treatment discontinuation. In addition, because clinical trials' participants usually represent non‐random samples from target populations, we describe two sensitivity analysis approaches. All discussions are motivated by an analysis that aims to characterize the dynamic relationship between concentrations of a liver enzyme (alanine aminotransferase) and three distinct doses (no drug, low dose, and high dose) of an nk‐1 antagonist across four Phase II clinical trials. Copyright © 2007 John Wiley & Sons, Ltd.
Penile fracture: diagnosis and management
Authors: Masarani M. , Dinneen M. 
Published: October 2007

AbstractPenile fracture is an uncommon urological emergency that can have significant complications. Patients usually present to the accident and emergency department, but may seek advice from GPs. Immediate surgical repair is the treatment of choice, with fewer complications, shorter hospital stays, better outcome and increased patient satisfaction compared with conservative therapy. Copyright © 2007 Wiley Interface Ltd