Browsing by Author "Shang, Barry Z."

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Kinetic Modeling at Single-Molecule Resolution Elucidates the Mechanisms of Cellulase Synergy
(2014-07-01)
Shang, Barry Z.
;
Chu, Jhih-Wei
;
生物科技學系
;
生物資訊及系統生物研究所
;
Department of Biological Science and Technology
The mechanism of synergistic cooperation between celluloses in decomposing cellulose is revealed by resolving the molecular structures of enzymes and substrates via kinetic modeling. The emergence of endo-exo and exo-exo synergy through enzyme-enzyme and enzyme-substrate couplings was investigated with the main exoglucanases (TrCel7A and TrCel6A) and endoglucanase (TrCel7B) of the Trichoderma reesei fungus. The degree of synergy was found to depend on the interplay between two competing effects: (1) enhancement of the complexation rates of exoglucanases with the chain ends created by endoglucanases and (2) surface inhibition of processive exoglucanases by uneven layers of glucan chains on cellulose. We show that the sole effect of TrCel7B in creating more free ends for TrCel7A is insufficient to cause synergistic activity. The combined actions of TrCel7A and TrCel7B produce a rougher substrate surface that subsequently promotes blocking of processing TrCel7A enzymes. This anti-synergy can be counteracted by enhancing the rates of complexation of TrCel7A with the TrCel7B-created chain ends, allowing their synergistic cooperation. Similarly, kinetic simulations show that exo-exo synergy does not occur if TrCel6A and TrCel7A have only opposite specificities in targeting the two ends of glucan chains. Incorporating endo activity into TrCel6A with complexation rate enhancement, however, can lead to synergy with TrCel7A. Therefore, we find that endo-exo and exo-exo synergies may share the same mechanistic origin. The results of work also highlight that resolving molecular configurations in kinetic modeling allows systematic analysis for elucidating the mechanism of interfacial biocatalysis.
Kinetic Modeling at Single-Molecule Resolution Elucidates the Mechanisms of Cellulase Synergy
(2014-07-01)
Shang, Barry Z.
;
Chu, Jhih-Wei
;
生物科技學系
;
生物資訊及系統生物研究所
;
Department of Biological Science and Technology
The mechanism of synergistic cooperation between celluloses in decomposing cellulose is revealed by resolving the molecular structures of enzymes and substrates via kinetic modeling. The emergence of endo-exo and exo-exo synergy through enzyme-enzyme and enzyme-substrate couplings was investigated with the main exoglucanases (TrCel7A and TrCel6A) and endoglucanase (TrCel7B) of the Trichoderma reesei fungus. The degree of synergy was found to depend on the interplay between two competing effects: (1) enhancement of the complexation rates of exoglucanases with the chain ends created by endoglucanases and (2) surface inhibition of processive exoglucanases by uneven layers of glucan chains on cellulose. We show that the sole effect of TrCel7B in creating more free ends for TrCel7A is insufficient to cause synergistic activity. The combined actions of TrCel7A and TrCel7B produce a rougher substrate surface that subsequently promotes blocking of processing TrCel7A enzymes. This anti-synergy can be counteracted by enhancing the rates of complexation of TrCel7A with the TrCel7B-created chain ends, allowing their synergistic cooperation. Similarly, kinetic simulations show that exo-exo synergy does not occur if TrCel6A and TrCel7A have only opposite specificities in targeting the two ends of glucan chains. Incorporating endo activity into TrCel6A with complexation rate enhancement, however, can lead to synergy with TrCel7A. Therefore, we find that endo-exo and exo-exo synergies may share the same mechanistic origin. The results of work also highlight that resolving molecular configurations in kinetic modeling allows systematic analysis for elucidating the mechanism of interfacial biocatalysis.
Linking hydrophobicity and hydrodynamics by the hybrid fluctuating hydrodynamics and molecular dynamics methodologies
(2013-08-27)
Voulgarakis, Nikolaos K.
;
Shang, Barry Z.
;
Chu, Jhih-Wei
;
生物科技學系
;
生物資訊及系統生物研究所
The development of a hybrid fluctuating hydrodynamics (FHD) and molecular dynamics (MD) simulation method that combines the molecular dynamics of moving particles with the fluctuating hydrodynamics of solvent fields on Eulerian grid cells is presented. This method allows resolution of solute-solvent interfaces and realization of excluded volumes of particles in the presence of hydrodynamic coupling. With these capabilities, we show that the ubiquitous forces mediated by the solvent, hydrophobicity and hydrodynamics, can be linked in a mesoscopic simulation. The strategies we devise to overcome the numerical issues of mixing variables in the Eulerian and Lagrangian coordinate systems, i.e., using a pair of auxiliary fluids to realize the excluded volumes of particles and assigning collocating gridding systems on solutes to interface with solvent fields, are also presented. Simulation results show that the hybrid FHD and MD method can reproduce the solvation free energies and scaling laws of particles dynamics for hydrophobes of different sizes. The collapse of two hydrophobic particles was also simulated to illustrate that the hybrid FHD and MD method has the potential to be generally applied to study nanoscale self-assembly and dynamics-structure-function relationships of biomolecules.
Systems-level Modeling with Molecular Resolution Elucidates the Rate-limiting Mechanisms of Cellulose Decomposition by Cellobiohydrolases
(2013-10-04)
Shang, Barry Z.
;
Chang, Rakwoo
;
Chu, Jhih-Wei
;
生物科技學系
;
生物資訊及系統生物研究所
Interprotein and enzyme-substrate couplings in interfacial biocatalysis induce spatial correlations beyond the capabilities of classical mass-action principles in modeling reaction kinetics. To understand the impact of spatial constraints on enzyme kinetics, we developed a computational scheme to simulate the reaction network of enzymes with the structures of individual proteins and substrate molecules explicitly resolved in the three-dimensional space. This methodology was applied to elucidate the rate-limiting mechanisms of crystalline cellulose decomposition by cellobiohydrolases. We illustrate that the primary bottlenecks are slow complexation of glucan chains into the enzyme active site and excessive enzyme jamming along the crowded substrate. Jamming could be alleviated by increasing the decomplexation rate constant but at the expense of reduced processivity. We demonstrate that enhancing the apparent reaction rate required a subtle balance between accelerating the complexation driving force and simultaneously avoiding enzyme jamming. Via a spatiotemporal systems analysis, we developed a unified mechanistic framework that delineates the experimental conditions under which different sets of rate-limiting behaviors emerge. We found that optimization of the complexation-exchange kinetics is critical for overcoming the barriers imposed by interfacial confinement and accelerating the apparent rate of enzymatic cellulose decomposition.