The Experts below are selected from a list of 585504 Experts worldwide ranked by ideXlab platform
Hsin-wei Chen - One of the best experts on this subject based on the ideXlab platform.
-
Immunogenicity of a novel tetravalent vaccine formulation with four recombinant lipidated dengue envelope Protein Domain IIIs in mice
Scientific Reports, 2016Co-Authors: Chen-yi Chiang, Chien-hsiung Pan, Mei-yu Chen, Chun-hsiang Hsieh, Jy-ping Tsai, Hsueh-hung Liu, Shih-jen Liu, Pele Chong, Chih-hsiang Leng, Hsin-wei ChenAbstract:We developed a novel platform to express high levels of recombinant lipoProteins with intrinsic adjuvant properties. Based on this technology, our group developed recombinant lipidated dengue envelope Protein Domain IIIs as vaccine candidates against dengue virus. This work aims to evaluate the immune responses in mice to the tetravalent formulation. We demonstrate that 4 serotypes of recombinant lipidated dengue envelope Protein Domain III induced both humoral and cellular immunity against all 4 serotypes of dengue virus on the mixture that formed the tetravalent formulation. Importantly, the immune responses induced by the tetravalent formulation in the absence of the exogenous adjuvant were functional in clearing the 4 serotypes of dengue virus in vivo . We affirm that the tetravalent formulation of recombinant lipidated dengue envelope Protein Domain III is a potential vaccine candidate against dengue virus and suggest further detailed studies of this formulation in nonhuman primates.
-
the immunodominance change and protection of cd4 t cell responses elicited by an envelope Protein Domain iii based tetravalent dengue vaccine in mice
PLOS ONE, 2015Co-Authors: Chen-yi Chiang, Chun-hsiang Hsieh, Hsin-wei Chen, Huimei Hu, Szuhsien Wu, Yuju Hsiao, Chiakai Wu, Hanhsuan Chung, Pele ChongAbstract:Dengue is the leading cause of mosquito-borne viral infections and no vaccine is available now. Envelope Protein Domain III (ED3) is the major target for the binding of dengue virus neutralizing antibodies; however, the ED3-specifc T-cell response is less well understood. To investigate the T-cell responses to four serotypes of dengue virus (DENV-1 to 4), we immunized mice using either a tetravalent ED3-based DNA or Protein vaccine, or combined both as a DNA prime-Protein boost strategy (prime-boost). A significant serotype-dependent IFN-γ or IL-4 response was observed in mice immunized with either the DNA or Protein vaccine. The IFN-γ response was dominant to DENV-1 to 3, whereas the IL-4 response was dominant to DENV-4. Although the similar IgG titers for the four serotypes were observed in mice immunized with the tetravalent vaccines, the neutralizing antibody titers varied and followed the order of 2 = 3>1>4. Interestingly, the lower IFN-γ response to DENV-4 is attributable to the immunodominance change between two CD4+ T-cell epitopes; one T-cell epitope located at E349-363 of DENV-1 to 3 was more immunogenic than the DENV-4 epitope E313-327. Despite DENV-4 specific IFN-γ responses were suppressed by immunodominance change, either DENV-4-specific IFN-γ or neutralizing antibody responses were still recalled after DENV-4 challenge and contributed to virus clearance. Immunization with the prime-boost elicited both IFN-γ and neutralizing antibody responses and provided better protection than either DNA or Protein immunization. Our findings shed light on how ED3-based tetravalent dengue vaccines sharpen host CD4 T-cell responses and contribute to protection against dengue virus.
David J E Callaway - One of the best experts on this subject based on the ideXlab platform.
-
nanoscale Protein Domain motion and long range allostery in signaling Proteins a view from neutron spin echo spectroscopy
Biophysical Reviews, 2015Co-Authors: David J E Callaway, Zimei BuAbstract:Many cellular Proteins are multi-Domain Proteins. Coupled Domain–Domain interactions in these multiDomain Proteins are important for the allosteric relay of signals in the cellular signaling networks. We have initiated the application of neutron spin echo spectroscopy to the study of nanoscale Protein Domain motions on submicrosecond time scales and on nanometer length scale. Our NSE experiments reveal the activation of Protein Domain motions over a long distance of over more than 100 A in a multiDomain scaffolding Protein NHERF1 upon binding to another Protein, Ezrin. Such activation of nanoscale Protein Domain motions is correlated with the allosteric assembly of multi-Protein complexes by NHERF1 and Ezrin. Here, we summarize the theoretical framework that we have developed, which uses simple concepts from nonequilibrium statistical mechanics to interpret the NSE data, and employs a mobility tensor to describe nanoscale Protein Domain motion. Extracting nanoscale Protein Domain motion from the NSE does not require elaborate molecular dynamics simulations, nor complex fits to rotational motion, nor elastic network models. The approach is thus more robust than multiparameter techniques that require untestable assumptions. We also demonstrate that an experimental scheme of selective deuteration of a Protein subunit in a complex can highlight and amplify specific Domain dynamics from the abundant global translational and rotational motions in a Protein. We expect NSE to provide a unique tool to determine nanoscale Protein dynamics for the understanding of Protein functions, such as how signals are propagated in a Protein over a long distance to a distal Domain.
-
coupled Protein Domain motion in taq polymerase revealed by neutron spin echo spectroscopy
Proceedings of the National Academy of Sciences of the United States of America, 2005Co-Authors: Zimei Bu, Ralf Biehl, M Monkenbusch, D Richter, David J E CallawayAbstract:Long-range conformational changes in Proteins are ubiquitous in biology for the transmission and amplification of signals; such conformational changes can be triggered by small-amplitude, nanosecond Protein Domain motion. Understanding how conformational changes are initiated requires the characterization of Protein Domain motion on these timescales and on length scales comparable to Protein dimensions. Using neutron spin-echo spectroscopy (NSE), normal mode analysis, and a statistical-mechanical framework, we reveal overdamped, coupled Domain motion within DNA polymerase I from Thermus aquaticus (Taq polymerase). This Protein utilizes correlated Domain dynamics over 70 A to coordinate nucleotide synthesis and cleavage during DNA synthesis and repair. We show that NSE spectroscopy can determine the Domain mobility tensor, which determines the degree of dynamical coupling between Domains. The mobility tensor defines the Domain velocity response to a force applied to it or to another Domain, just as the sails of a sailboat determine its velocity given the applied wind force. The NSE results provide insights into the nature of Protein Domain motion that are not appreciated by conventional biophysical techniques.
Zimei Bu - One of the best experts on this subject based on the ideXlab platform.
-
nanoscale Protein Domain motion and long range allostery in signaling Proteins a view from neutron spin echo spectroscopy
Biophysical Reviews, 2015Co-Authors: David J E Callaway, Zimei BuAbstract:Many cellular Proteins are multi-Domain Proteins. Coupled Domain–Domain interactions in these multiDomain Proteins are important for the allosteric relay of signals in the cellular signaling networks. We have initiated the application of neutron spin echo spectroscopy to the study of nanoscale Protein Domain motions on submicrosecond time scales and on nanometer length scale. Our NSE experiments reveal the activation of Protein Domain motions over a long distance of over more than 100 A in a multiDomain scaffolding Protein NHERF1 upon binding to another Protein, Ezrin. Such activation of nanoscale Protein Domain motions is correlated with the allosteric assembly of multi-Protein complexes by NHERF1 and Ezrin. Here, we summarize the theoretical framework that we have developed, which uses simple concepts from nonequilibrium statistical mechanics to interpret the NSE data, and employs a mobility tensor to describe nanoscale Protein Domain motion. Extracting nanoscale Protein Domain motion from the NSE does not require elaborate molecular dynamics simulations, nor complex fits to rotational motion, nor elastic network models. The approach is thus more robust than multiparameter techniques that require untestable assumptions. We also demonstrate that an experimental scheme of selective deuteration of a Protein subunit in a complex can highlight and amplify specific Domain dynamics from the abundant global translational and rotational motions in a Protein. We expect NSE to provide a unique tool to determine nanoscale Protein dynamics for the understanding of Protein functions, such as how signals are propagated in a Protein over a long distance to a distal Domain.
-
coupled Protein Domain motion in taq polymerase revealed by neutron spin echo spectroscopy
Proceedings of the National Academy of Sciences of the United States of America, 2005Co-Authors: Zimei Bu, Ralf Biehl, M Monkenbusch, D Richter, David J E CallawayAbstract:Long-range conformational changes in Proteins are ubiquitous in biology for the transmission and amplification of signals; such conformational changes can be triggered by small-amplitude, nanosecond Protein Domain motion. Understanding how conformational changes are initiated requires the characterization of Protein Domain motion on these timescales and on length scales comparable to Protein dimensions. Using neutron spin-echo spectroscopy (NSE), normal mode analysis, and a statistical-mechanical framework, we reveal overdamped, coupled Domain motion within DNA polymerase I from Thermus aquaticus (Taq polymerase). This Protein utilizes correlated Domain dynamics over 70 A to coordinate nucleotide synthesis and cleavage during DNA synthesis and repair. We show that NSE spectroscopy can determine the Domain mobility tensor, which determines the degree of dynamical coupling between Domains. The mobility tensor defines the Domain velocity response to a force applied to it or to another Domain, just as the sails of a sailboat determine its velocity given the applied wind force. The NSE results provide insights into the nature of Protein Domain motion that are not appreciated by conventional biophysical techniques.
Eugene I Shakhnovich - One of the best experts on this subject based on the ideXlab platform.
-
a simple model of Protein Domain swapping in crowded cellular environments
Biophysical Journal, 2016Co-Authors: Jaie C Woodard, Sachith Dunatunga, Eugene I ShakhnovichAbstract:Domain swapping in Proteins is an important mechanism of functional and structural innovation. However, despite its ubiquity and importance, the physical mechanisms that lead to Domain swapping are poorly understood. Here, we present a simple two-dimensional coarse-grained model of Protein Domain swapping in the cytoplasm. In our model, two-Domain Proteins partially unfold and diffuse in continuous space. Monte Carlo multiProtein simulations of the model reveal that Domain swapping occurs at intermediate temperatures, whereas folded dimers and folded monomers prevail at low temperatures, and partially unfolded monomers predominate at high temperatures. We use a simplified amino acid alphabet consisting of four residue types, and find that the oligomeric state at a given temperature depends on the sequence of the Protein. We also show that hinge strain between Domains can promote Domain swapping, consistent with experimental observations for real Proteins. Domain swapping depends nonmonotonically on the Protein concentration, with Domain-swapped dimers occurring at intermediate concentrations and nonspecific interactions between partially unfolded Proteins occurring at high concentrations. For folded Proteins, we recover the result obtained in three-dimensional lattice simulations, i.e., that functional dimerization is most prevalent at intermediate temperatures and nonspecific interactions increase at low temperatures.
-
reconstruction of the src sh3 Protein Domain transition state ensemble using multiscale molecular dynamics simulations
Journal of Molecular Biology, 2005Co-Authors: Feng Ding, Nikolay V Dokholyan, Eugene I Shakhnovich, Joanemma SheaAbstract:We use an integrated computational approach to reconstruct accurately the transition state ensemble (TSE) for folding of the src-SH3 Protein Domain. We first identify putative TSE conformations from free energy surfaces generated by importance sampling molecular dynamics for a fully atomic, solvated model of the src-SH3 Protein Domain. These putative TSE conformations are then subjected to a folding analysis using a coarse-grained representation of the Protein and rapid discrete molecular dynamics simulations. Those conformations that fold to the native conformation with a probability (Pfold) of approximately 0.5, constitute the true transition state. Approximately 20% of the putative TSE structures were found to have a Pfold near 0.5, indicating that, although correct TSE conformations are populated at the free energy barrier, there is a critical need to refine this ensemble. Our simulations indicate that the true TSE conformations are compact, with a well-defined central β sheet, in good agreement with previous experimental and theoretical studies. A structured central β sheet was found to be present in a number of pre-TSE conformations, however, indicating that this element, although required in the transition state, does not define it uniquely. An additional tight cluster of contacts between highly conserved residues belonging to the diverging turn and second β-sheet of the Protein emerged as being critical elements of the folding nucleus. A number of commonly used order parameters to identify the transition state for folding were investigated, with the number of native Cβ contacts displaying the most satisfactory correlation with Pfold values.
-
folding kinetics of villin 14t a Protein Domain with a central beta sheet and two hydrophobic cores
Biochemistry, 1998Co-Authors: Sung E Choe, Paul Matsudaira, John J Osterhout, Gerhard Wagner, Eugene I ShakhnovichAbstract:The thermodynamics and kinetics of folding are characterized for villin 14T, a 126-residue Protein Domain. Equilibrium fluorescence measurements reveal that villin 14T unfolds and refolds reversibly. The folding kinetics was monitored using stopped-flow with fluorescence and quenched-flow with NMR and mass spectrometry. Unfolding occurs in a single-exponential phase in the stopped-flow experiments, and about 75% of the total amplitude is recovered in the fast phase of refolding. The remaining 25% of the amplitude probably represents trapping in cis−trans proline isomerization pathways. At 25 °C, the stability estimate obtained by extrapolation from the transition region of the stopped-flow chevron matches the stability value from equilibrium urea titrations (ΔG = 9.7 kcal/mol, m value = 2.2 kcal mol-1 M-1). At low final urea concentrations, however, the refolding kinetics deviates from the two-state model, indicating the formation of an intermediate. Under these conditions, quenched-flow followed by NMR a...
Joanemma Shea - One of the best experts on this subject based on the ideXlab platform.
-
reconstruction of the src sh3 Protein Domain transition state ensemble using multiscale molecular dynamics simulations
Journal of Molecular Biology, 2005Co-Authors: Feng Ding, Nikolay V Dokholyan, Eugene I Shakhnovich, Joanemma SheaAbstract:We use an integrated computational approach to reconstruct accurately the transition state ensemble (TSE) for folding of the src-SH3 Protein Domain. We first identify putative TSE conformations from free energy surfaces generated by importance sampling molecular dynamics for a fully atomic, solvated model of the src-SH3 Protein Domain. These putative TSE conformations are then subjected to a folding analysis using a coarse-grained representation of the Protein and rapid discrete molecular dynamics simulations. Those conformations that fold to the native conformation with a probability (Pfold) of approximately 0.5, constitute the true transition state. Approximately 20% of the putative TSE structures were found to have a Pfold near 0.5, indicating that, although correct TSE conformations are populated at the free energy barrier, there is a critical need to refine this ensemble. Our simulations indicate that the true TSE conformations are compact, with a well-defined central β sheet, in good agreement with previous experimental and theoretical studies. A structured central β sheet was found to be present in a number of pre-TSE conformations, however, indicating that this element, although required in the transition state, does not define it uniquely. An additional tight cluster of contacts between highly conserved residues belonging to the diverging turn and second β-sheet of the Protein emerged as being critical elements of the folding nucleus. A number of commonly used order parameters to identify the transition state for folding were investigated, with the number of native Cβ contacts displaying the most satisfactory correlation with Pfold values.
-
probing the folding free energy landscape of the src sh3 Protein Domain
Proceedings of the National Academy of Sciences of the United States of America, 2002Co-Authors: Joanemma Shea, Jose N Onuchic, Charles L BrooksAbstract:The mechanism and thermodynamics of folding of the Src homology 3 (SH3) Protein Domain are characterized at an atomic level through molecular dynamics with importance sampling. This methodology enables the construction of the folding free energy landscape of the Protein as a function of representative reaction coordinates. We observe that folding proceeds in a downhill manner under native conditions, with early compaction and structure formation in the hydrophobic sheet consisting of the three central β strands of the Protein. This state bears considerable resemblance to the experimentally determined transition state for folding. Folding proceeds further with the formation of the second hydrophobic sheet consisting of the terminal strands and the RT loop. The final stages of folding appear to involve the formation of the hydrophobic core through the expulsion of water molecules bridging the two hydrophobic sheets. This work sheds new light on the complementary roles of sequence and topology in governing the folding mechanism of small Proteins and provides further support for the role of water in facilitating the late stages in folding.
