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Nucleic Acids Research Jul 2017Antibodies have become an indispensable tool for many biotechnological and clinical applications. They bind their molecular target (antigen) by recognizing a portion of...
Antibodies have become an indispensable tool for many biotechnological and clinical applications. They bind their molecular target (antigen) by recognizing a portion of its structure (epitope) in a highly specific manner. The ability to predict epitopes from antigen sequences alone is a complex task. Despite substantial effort, limited advancement has been achieved over the last decade in the accuracy of epitope prediction methods, especially for those that rely on the sequence of the antigen only. Here, we present BepiPred-2.0 (http://www.cbs.dtu.dk/services/BepiPred/), a web server for predicting B-cell epitopes from antigen sequences. BepiPred-2.0 is based on a random forest algorithm trained on epitopes annotated from antibody-antigen protein structures. This new method was found to outperform other available tools for sequence-based epitope prediction both on epitope data derived from solved 3D structures, and on a large collection of linear epitopes downloaded from the IEDB database. The method displays results in a user-friendly and informative way, both for computer-savvy and non-expert users. We believe that BepiPred-2.0 will be a valuable tool for the bioinformatics and immunology community.
Topics: Epitopes, B-Lymphocyte; Internet; Models, Molecular; Muramidase; Protein Conformation; Sequence Analysis, Protein; Software; User-Computer Interface
PubMed: 28472356
DOI: 10.1093/nar/gkx346 -
Langmuir : the ACS Journal of Surfaces... Apr 2019We discuss recent investigations of the interaction of polyelectrolytes with proteins. In particular, we review our recent studies on the interaction of simple proteins... (Review)
Review
We discuss recent investigations of the interaction of polyelectrolytes with proteins. In particular, we review our recent studies on the interaction of simple proteins such as human serum albumin (HSA) and lysozyme with linear polyelectrolytes, charged dendrimers, charged networks, and polyelectrolyte brushes. In all cases discussed here, we combined experimental work with molecular dynamics (MD) simulations and mean-field theories. In particular, isothermal titration calorimetry (ITC) has been employed to obtain the respective binding constants K and the Gibbs free energy of binding. MD simulations with explicit counterions but implicit water demonstrate that counterion release is the main driving force for the binding of proteins to strongly charged polyelectrolytes: patches of positive charges located on the surface of the protein become multivalent counterions of the polyelectrolyte, thereby releasing a number of counterions condensed on the polyelectrolyte. The binding Gibbs free energy due to counterion release is predicted to scale with the logarithm of the salt concentration in the system, which is verified by both simulations and experiment. In several cases, namely, for the interaction of proteins with linear polyelectrolytes and highly charged hydrophilic dendrimers, the binding constant could be calculated from simulations to very good approximation. This finding demonstrated that in these cases explicit hydration effects do not contribute to the Gibbs free energy of binding. The Gibbs free energy can also be used to predict the kinetics of protein uptake by microgels for a given system by applying dynamic density functional theory. The entire discussion demonstrates that the direct comparison of theory with experiments can lead to a full understanding of the interaction of proteins with charged polymers. Possible implications for applications, such as drug design, are discussed.
Topics: Humans; Molecular Dynamics Simulation; Muramidase; Polyelectrolytes; Protein Binding; Serum Albumin, Human; Thermodynamics
PubMed: 30095921
DOI: 10.1021/acs.langmuir.8b01802 -
Journal of Computational Chemistry Jun 2021Computation of the thermodynamic consequences of protein mutations holds great promise in protein biophysics and design. Alchemical free energy methods can give improved...
Computation of the thermodynamic consequences of protein mutations holds great promise in protein biophysics and design. Alchemical free energy methods can give improved estimates of mutational free energies, and are already widely used in calculations of relative and absolute binding free energies in small molecule design problems. In principle, alchemical methods can address any amino acid mutation with an appropriate alchemical pathway, but identifying a strategy that produces such a path for proline and glycine mutations is an ongoing challenge. Most current strategies perturb only side chain atoms, while proline and glycine mutations also alter the backbone parameters and backbone ring topology. Some strategies also perturb backbone parameters and enable glycine mutations. This work presents a strategy that enables both proline and glycine mutations and comprises two key elements: a dual backbone with restraints and scaling of bonded terms, facilitating backbone parameter changes, and a soft bond in the proline ring, enabling ring topology changes in proline mutations. These elements also have utility for core hopping and macrocycle studies in computer-aided drug design. This new strategy shows slight improvements over an alternative side chain perturbation strategy for a set T4 lysozyme mutations lacking proline and glycine, and yields good agreement with experiment for a set of T4 lysozyme proline and glycine mutations not previously studied. To our knowledge this is the first report comparing alchemical predictions of proline mutations with experiment. With this strategy in hand, alchemical methods now have access to the full palette of amino acid mutations.
Topics: Glycine; Molecular Dynamics Simulation; Muramidase; Mutation; Proline; Thermodynamics
PubMed: 33844328
DOI: 10.1002/jcc.26525 -
Acta Crystallographica. Section D,... Jan 2022The accuracy of B factors in protein crystal structures has been determined by comparing the same atoms in numerous, independent crystal structures of Gallus gallus...
The accuracy of B factors in protein crystal structures has been determined by comparing the same atoms in numerous, independent crystal structures of Gallus gallus lysozyme. Both B-factor absolute differences and normal probability plots indicate that the estimated B-factor errors are quite large, close to 9 Å in ambient-temperature structures and to 6 Å in low-temperature structures, and surprisingly are comparable to values estimated two decades ago. It is well known that B factors are not due to local movements only but reflect several, additional factors from crystal defects, large-scale disorder, diffraction data quality etc. It therefore remains essential to normalize B factors when comparing different crystal structures, although it has clearly been shown that they provide useful information about protein dynamics. Improved, quantitative analyses of raw B factors require novel experimental and computational tools that are able to disaggregate local movements from other features and properties that affect B factors.
Topics: Algorithms; Animals; Chickens; Computational Biology; Crystallization; Crystallography, X-Ray; Molecular Structure; Muramidase; Protein Conformation; Reproducibility of Results; Temperature; X-Ray Diffraction
PubMed: 34981763
DOI: 10.1107/S2059798321011736 -
Indian Journal of Dermatology,... 2017Autophagy, literally meaning "self-eating," is an intracellular catabolic process of delivering cytosol and/or its specific content to the lysosomes for degradation.The... (Review)
Review
Autophagy, literally meaning "self-eating," is an intracellular catabolic process of delivering cytosol and/or its specific content to the lysosomes for degradation.The resulting macromolecular constituents are recycled and utilized again by the cells. Basal level autophagy plays an important role in cellular homeostasis through the elimination of the old or damaged organelles, as well as aggregated intracellular proteins. Autophagy refers to sequestration of intact organelles along with a portion of cytosol, into a double-or multi-membrane structure known as phagophore, which elongates, and after closure, forms a vesicular structure known as the autophagosome. Subsequently, the mature autophagosome fuses with a lysosome, thereby forming a single membrane structure, an autolysosome. Autophagy plays a critical role in inflammation, autoimmunity and cellular differentiation. Skin serves as the first line of defense against a variety of environmental insults and autophagy is thought to be a form of an endogenous defense mechanism against such environmental derangements. Autophagy has been linked with keratinocyte differentiation and melanocyte survival, as well as with the pathogenesis of diverse skin disorders including systemic lupus erythematosus, systemic sclerosis, psoriasis, vitiligo, infectious skin diseases and cancer. Autophagy has been one of the most studied phenomena in cell biology and pathophysiology, and given its broad clinical implications, has become a major target for drug discovery. The last decade has seen a substantial upsurge in autophagy-related research and publications; still, the dermatology literature appears to be less initiated. Autophagy will probably change our understanding of dermatological disorders/medicines. Hence, a basic knowledge of autophagy is a prerequisite to understand the developments in the field of autophagy-related research.
Topics: Animals; Autophagy; Dermatologic Agents; Dermatology; Humans; Muramidase; Protein Kinases; Skin Diseases
PubMed: 28004651
DOI: 10.4103/0378-6323.196320 -
BMC Evolutionary Biology Aug 2014Lysozyme g is an antibacterial enzyme that was first found in the eggs of some birds, but recently has been found in additional species, including non-vertebrates. Some...
BACKGROUND
Lysozyme g is an antibacterial enzyme that was first found in the eggs of some birds, but recently has been found in additional species, including non-vertebrates. Some previously characterized lysozyme g sequences are suggested to have altered secretion potential and enzymatic activity, however the distribution of these altered sequences is unknown. Duplicated copies of the lysozyme g gene exist in some species; however, the origins of the duplicates and their roles in altered function are unclear.
RESULTS
We identified 234 lysozyme g sequences from 118 vertebrate species, including 181 sequences that are full or near full length representing all vertebrate classes except cartilaginous fish. Phylogenetic analysis shows that most lysozyme g gene duplicates are recent or lineage specific events, however three amplification events are more ancient, those in an early amniote, an early mammal, and an early teleost. The older gene duplications are associated with changes in function, including changes in secretion potential and muramidase antibacterial enzymatic activity.
CONCLUSIONS
Lysozyme g is an essential muramidase enzyme that is widespread in vertebrates. Duplication of the lysozyme g gene, and the retention of non-secreted isozymes that have lost enzymatic activity indicate that lysozyme g has an activity other than the muramidase activity associated with being an antibacterial enzyme.
Topics: Animals; Base Sequence; Evolution, Molecular; Geese; Gene Duplication; Multigene Family; Muramidase; Phylogeny
PubMed: 25167808
DOI: 10.1186/s12862-014-0188-x -
Nature Communications Oct 2021Nanopores are single-molecule sensors used in nucleic acid analysis, whereas their applicability towards full protein identification has yet to be demonstrated. Here, we...
Nanopores are single-molecule sensors used in nucleic acid analysis, whereas their applicability towards full protein identification has yet to be demonstrated. Here, we show that an engineered Fragaceatoxin C nanopore is capable of identifying individual proteins by measuring peptide spectra that are produced from hydrolyzed proteins. Using model proteins, we show that the spectra resulting from nanopore experiments and mass spectrometry share similar profiles, hence allowing protein fingerprinting. The intensity of individual peaks provides information on the concentration of individual peptides, indicating that this approach is quantitative. Our work shows the potential of a low-cost, portable nanopore-based analyzer for protein identification.
Topics: Calibration; Cnidarian Venoms; Hydrolysis; Muramidase; Nanopores; Peptide Mapping; Peptides; Proteins
PubMed: 34608150
DOI: 10.1038/s41467-021-26046-9 -
Quarterly Reviews of Biophysics Feb 2012Molecular recognition plays a central role in biochemical processes. Although well studied, understanding the mechanisms of recognition is inherently difficult due to... (Review)
Review
Molecular recognition plays a central role in biochemical processes. Although well studied, understanding the mechanisms of recognition is inherently difficult due to the range of potential interactions, the molecular rearrangement associated with binding, and the time and length scales involved. Computational methods have the potential for not only complementing experiments that have been performed, but also in guiding future ones through their predictive abilities. In this review, we discuss how molecular dynamics (MD) simulations may be used in advancing our understanding of the thermodynamics that drive biomolecular recognition. We begin with a brief review of the statistical mechanics that form a basis for these methods. This is followed by a description of some of the most commonly used methods: thermodynamic pathways employing alchemical transformations and potential of mean force calculations, along with end-point calculations for free energy differences, and harmonic and quasi-harmonic analysis for entropic calculations. Finally, a few of the fundamental findings that have resulted from these methods are discussed, such as the role of configurational entropy and solvent in intermolecular interactions, along with selected results of the model system T4 lysozyme to illustrate potential and current limitations of these methods.
Topics: Biochemistry; Endpoint Determination; Entropy; Molecular Dynamics Simulation; Muramidase; Probability Theory
PubMed: 22082669
DOI: 10.1017/S0033583511000096 -
Postepy Higieny I Medycyny... Dec 2014Lysozyme (LZ, muramidase, N-acetylmuramylhydrolase) is a protein occuring in animals, plants, bacteria and viruses. It can be found e.g. in granules of neutrophils,... (Review)
Review
Lysozyme (LZ, muramidase, N-acetylmuramylhydrolase) is a protein occuring in animals, plants, bacteria and viruses. It can be found e.g. in granules of neutrophils, macrophages and in serum, saliva, milk, honey and hen egg white. The enzyme hydrolyzes the β-1,4 glycosidic bonds between N-acetylmuramic acid (NAM) and N-acetylglucosamine (NAG) of cell wall peptidoglycan (PG) in Gram-positive and Gram-negative bacteria. In the animal kingdom, three muramidase types have been identified: the c-type (chicken type), the g-type (goose-type) and the i-type (invertebrates). The c-type LZ from hen egg white is a model for the study of protein structure and function. Muramidase shows bactericidal activity mainly against Gram-positive bacteria. Cytolytic activity against cells of Gram-negative bacteria has not been proved. Bacterial cells have developed defense mechanisms that allow them to avoid the action of LZ. They are based e.g. on the production of enzyme inhibitors or modification of the PG. LZ is one of the most studied enzymes and yet not all aspects characterizing this protein are fully understood. One of the most important unresolved issues concerning the biological function of LZ is the role of muramidase in the bactericidal action of serum against Gram-negative bacteria. In order to clarify the function of LZ, the enzyme is e.g. removed from the serum by adsorption onto bentonite (montmorillonite, MMT). By using X-ray diffraction techniques it has been shown that MMT after contact with the serum is delaminated. The problems associated with folding of muramidase and LZ participation in the development of amyloidoses also await explanation.
Topics: Animals; Anti-Bacterial Agents; Gram-Negative Bacteria; Gram-Positive Bacteria; Muramidase
PubMed: 25531714
DOI: 10.5604/17322693.1133100 -
Scientific Reports Feb 2022Macromolecular protein crystallisation was one of the potential tools to accelerate the biomanufacturing of biopharmaceuticals. In this work, it was the first time to...
Macromolecular protein crystallisation was one of the potential tools to accelerate the biomanufacturing of biopharmaceuticals. In this work, it was the first time to investigate the roles of biotemplates, Saccharomyces cerevisiae live cells, in the crystallisation processes of lysozyme, with different concentrations from 20 to 2.5 mg/mL lysozyme and different concentrations from 0 to 5.0 × 10 (cfu/mL) Saccharomyces cerevisiae cells, during a period of 96 h. During the crystallisation period, the nucleation possibility in droplets, crystal numbers, and cell growth and cell density were observed and analysed. The results indicated the strong interaction between the lysozyme molecules and the cell wall of the S. cerevisiae, proved by the crystallization of lysozyme with fluorescent labels. The biotemplates demonstrated positive influence or negative influence on the nucleation, i.e. shorter or longer induction time, dependent on the concentrations of the lysozyme and the S. cerevisiae cells, and ratios between them. In the biomanufacturing process, target proteins were various cells were commonly mixed with various cells, and this work provides novel insights of new design and application of live cells as biotemplates for purification of macromolecules.
Topics: Biopharmaceutics; Cell Wall; Crystallization; Fluorescent Dyes; Macromolecular Substances; Muramidase; Saccharomyces cerevisiae
PubMed: 35194113
DOI: 10.1038/s41598-022-06999-7