Conserved conformational hierarchy across functionally divergent glycosyltransferases of the GT-B structural superfamily as determined from microsecond molecular dynamics

AbstractIt has long been understood that some proteins to undergo conformational transitions enroute to the Michaelis Complex to allow chemistry. Examination of crystal structures of glycosyltransferase enzymes in the GT-B structural class reveals that the presence of ligand in the active site is necessary for the protein to crystalize in the closed conformation. Herein we describe microsecond molecular dynamics simulations of two evolutionarily unrelated glycosytransferases, HepI and GtfA. Simulations were performed using these proteins in the open and closed conformations, (respectively,) and we sought to identify the major dynamical modes and communication networks which allow conformational transition between the open and closed structures. We provide the first reported evidence (within the scope of our experimental parameters) that conformational hierarchy/directionality of the interconversion between open and closed conformations is a conserved feature of enzymes of the same structural superfamily. Additionally, residues previously identified to be important for substrate binding in HepI were shown to have strong negative correlations with non-ionizable residues distal to the active site. Mutagenesis of these residues produced mutants with altered enzymatic efficiency exhibiting lower Km values, while the kcat is effectively unchanged. The negatively correlated motions of these residues are important for substrate binding and forming the Michaelis complex, without impacting the activation barrier for catalysis. This suggests that in the bi-domain HepI, the enzyme dynamics did not impact the transition state stabilization or chemistry, but rather earlier steps along the reaction coordinate, leading to the reorganization of the active site electrostatic environment required for catalysis.

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Conserved Conformational Hierarchy across Functionally Divergent Glycosyltransferases of the GT-B Structural Superfamily as Determined from Microsecond Molecular Dynamics

International Journal of Molecular Sciences ◽

10.3390/ijms22094619 ◽

2021 ◽

Vol 22 (9) ◽

pp. 4619

Author(s):

Carlos A. Ramirez-Mondragon ◽

Megin E. Nguyen ◽

Jozafina Milicaj ◽

Bakar A. Hassan ◽

Frank J. Tucci ◽

...

Keyword(s):

Molecular Dynamics ◽

Communication Networks ◽

Active Site ◽

Conformational Dynamics ◽

Catalytic Efficiency ◽

Potential Effect ◽

Structural Class ◽

Terminal Domain ◽

Cross Correlation Analysis ◽

Dynamics Simulations

It has long been understood that some proteins undergo conformational transitions en route to the Michaelis Complex to allow chemistry. Examination of crystal structures of glycosyltransferase enzymes in the GT-B structural class reveals that the presence of ligand in the active site triggers an open-to-closed conformation transition, necessary for their catalytic functions. Herein, we describe microsecond molecular dynamics simulations of two distantly related glycosyltransferases that are part of the GT-B structural superfamily, HepI and GtfA. Simulations were performed using the open and closed conformations of these unbound proteins, respectively, and we sought to identify the major dynamical modes and communication networks that interconnect the open and closed structures. We provide the first reported evidence within the scope of our simulation parameters that the interconversion between open and closed conformations is a hierarchical multistep process which can be a conserved feature of enzymes of the same structural superfamily. Each of these motions involves of a collection of smaller molecular reorientations distributed across both domains, highlighting the complexities of protein dynamic involved in the interconversion process. Additionally, dynamic cross-correlation analysis was employed to explore the potential effect of distal residues on the catalytic efficiency of HepI. Multiple distal nonionizable residues of the C-terminal domain exhibit motions anticorrelated to positively charged residues in the active site in the N-terminal domain involved in substrate binding. Mutations of these residues resulted in a reduction in negatively correlated motions and an altered enzymatic efficiency that is dominated by lower Km values with kcat effectively unchanged. The findings suggest that residues with opposing conformational motions involved in the opening and closing of the bidomain HepI protein can allosterically alter the population and conformation of the “closed” state, essential to the formation of the Michaelis complex. The stabilization effects of these mutations likely equally influence the energetics of both the ground state and the transition state of the catalytic reaction, leading to the unaltered kcat. Our study provides new insights into the role of conformational dynamics in glycosyltransferase’s function and new modality to modulate enzymatic efficiency.

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Structural Differences In 3C-like protease (Mpro) From SARS-CoV and SARS-CoV-2: Molecular Insights For Drug Repurposing Against COVID-19 Revealed by Molecular Dynamics Simulations.

10.1101/2021.08.11.455903 ◽

2021 ◽

Author(s):

Dhaval Patel ◽

Meet Parmar ◽

Ritik Thumar ◽

Bhumi Patel ◽

Mohd. Athar ◽

...

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Binding Site ◽

Active Site ◽

Substrate Binding ◽

Drug Repurposing ◽

Molecular Networks ◽

Substrate Binding Site ◽

Sequence Identity ◽

Dynamics Simulations

A recent fatal outbreak of novel coronavirus SARS-CoV-2, identified preliminary as a causative agent for series of unusual pneumonia cases in Wuhan city, China has infected more than 20 million individuals with more than 4 million mortalities. Since, the infection crossed geographical barriers, the WHO permanently named the causing disease as COVID-2019 by declaring it a pandemic situation. SARS-CoV-2 is an enveloped single-stranded RNA virus causing a wide range of pathological conditions from common cold symptoms to pneumonia and fatal severe respiratory syndrome. Genome sequencing of SARS-CoV-2 has revealed 96% identity to the bat coronavirus and 79.6% sequence identity to the previous SARS-CoV. The main protease (known as 3C-like proteinase/ Mpro) plays a vital role during the infection with the processing of replicase polyprotein thus offering an attractive target for therapeutic interventions. SARS-CoV and SARS-CoV-2 Mpro shares 97% sequence identity, with 12 variable residues but none of them are present in the catalytic and substrate binding site. With the high level of sequence and structural similarity and absence of any drug/vaccine against SARS-CoV-2, drug repurposing against Mpro is an effective strategy to combat COVID-19. Here, we report a detailed comparison of SARS-CoV-2 Mpro with SARS-CoV Mpro using molecular dynamics simulations to assess the impact of 12 divergent residues on the molecular microenvironment of Mpro. Structural comparison and analysis are made on how these variable residues affect the intra-molecular interactions between key residues in the monomer and biologically active dimer form of Mpro. The present MD simulations study concluded the change in microenvironment of active-site residues at the entrance (T25, T26, M49 and Q189), near the catalytic region (F140, H163, H164, M165 and H172) and other residues in substrate binding site (V35T, N65S, K88R and N180K) due to 12 mutation incorporated in the SARS-CoV-2 Mpro. It is also evident that SARS-CoV-2 dimer is more stable and less flexible state compared to monomer which may be due to these variable residues, mainly F140, E166 and H172 which are involved in dimerization. This also warrants a need for inhibitor design considering the more stable dimer form. The mutation accumulated in SARS-CoV-2 Mpro indirectly reconfigures the key molecular networks around the active site conferring a potential change in SARS-CoV-2, thus posing a challenge in drug repurposing SARS drugs for COVID-19. The new networks and changes in the microenvironment identified by our work might guide attempts needed for repurposing and identification of new Mpro inhibitors.

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Conformation gating as a mechanism for enzyme specificity

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.95.16.9280 ◽

1998 ◽

Vol 95 (16) ◽

pp. 9280-9283 ◽

Cited By ~ 157

Author(s):

Huan-Xiang Zhou ◽

Stanislaw T. Wlodek ◽

J. Andrew McCammon

Keyword(s):

Molecular Dynamics ◽

Active Site ◽

Active Sites ◽

Substrate Binding ◽

Enzyme Specificity ◽

Aromatic Rings ◽

Binding Rate ◽

Molecular Dynamics Trajectory ◽

Dynamics Simulations ◽

Opening And Closing

Acetylcholinesterase, with an active site located at the bottom of a narrow and deep gorge, provides a striking example of enzymes with buried active sites. Recent molecular dynamics simulations showed that reorientation of five aromatic rings leads to rapid opening and closing of the gate to the active site. In the present study the molecular dynamics trajectory is used to quantitatively analyze the effect of the gate on the substrate binding rate constant. For a 2.4-Å probe modeling acetylcholine, the gate is open only 2.4% of the time, but the quantitative analysis reveals that the substrate binding rate is slowed by merely a factor of 2. We rationalize this result by noting that the substrate, by virtue of Brownian motion, will make repeated attempts to enter the gate each time it is near the gate. If the gate is rapidly switching between the open and closed states, one of these attempts will coincide with an open state, and then the substrate succeeds in entering the gate. However, there is a limit on the extent to which rapid gating dynamics can compensate for the small equilibrium probability of the open state. Thus the gate is effective in reducing the binding rate for a ligand 0.4 Å bulkier by three orders of magnitude. This relationship suggests a mechanism for achieving enzyme specificity without sacrificing efficiency.

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Role of Long-range Allosteric Communication in Determining the Stability and Disassembly of SARS-COV-2 in Complex with ACE2

10.1101/2020.11.30.405340 ◽

2020 ◽

Author(s):

Mauro L. Mugnai ◽

Clark Templeton ◽

Ron Elber ◽

D. Thirumalai

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Long Range ◽

Active Site ◽

Large Scale ◽

Conformational Transition ◽

Network Models ◽

Allosteric Communication ◽

The Stability ◽

Dynamics Simulations

AbstractSevere acute respiratory syndrome (SARS) and novel coronavirus disease (COVID-19) are caused by two closely related beta-coronaviruses, SARS-CoV and SARS-CoV-2, respectively. The envelopes surrounding these viruses are decorated with spike proteins, whose receptor binding domains (RBDs) initiate invasion by binding to the human angiotensin-converting enzyme 2 (ACE2). Subtle changes at the interface with ACE2 seem to be responsible for the enhanced affinity for the receptor of the SARS-CoV-2 RBD compared to SARS-CoV RBD. Here, we use Elastic Network Models (ENMs) to study the response of the viral RBDs and ACE2 upon dissassembly of the complexes. We identify a dominant detachment mode, in which the RBD rotates away from the surface of ACE2, while the receptor undergoes a conformational transition which stretches the active-site cleft. Using the Structural Perturbation Method, we determine the network of residues, referred to as the Allostery Wiring Diagram (AWD), which drives the large-scale motion activated by the detachment of the complex. The AWD for SARS-CoV and SARS-CoV-2 are remarkably similar, showing a network that spans the interface of the complex and reaches the active site of ACE2, thus establishing an allosteric connection between RBD binding and receptor catalytic function. Informed in part by the AWD, we used Molecular Dynamics simulations to probe the effect of interfacial mutations in which SARS-CoV-2 residues are replaced by their SARS-CoV counterparts. We focused on a conserved glycine (G502 in SARS-CoV-2, G488 in SARS-CoV) because it belongs to a region that initiates the dissociation of the complex along the dominant detachment mode, and is prominent in the AWD. Molecular Dynamics simulations of SARS-CoV-2 wild-type and G502P mutant show that the affinity for the human receptor of the mutant is drastically diminished. Our results suggest that in addition to residues that are in direct contact with the interface those involved in long range allosteric communication are also a determinant of the stability of the RBD-ACE2 complex.

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WHICH IS THE PROTON-SHUTTLE IN ISOXANTHOPTERIN DEAMINASE? QM/MM MD UNDERSTANDING

Journal of Theoretical and Computational Chemistry ◽

10.1142/s0219633613410022 ◽

2013 ◽

Vol 12 (08) ◽

pp. 1341002 ◽

Cited By ~ 1

Author(s):

XIN ZHANG ◽

MING LEI

Keyword(s):

Crystal Structure ◽

Molecular Dynamics ◽

Hydrogen Bonds ◽

Proton Transfer ◽

Glutamic Acid ◽

Active Site ◽

Nucleophilic Attack ◽

Zinc Ions ◽

Initial Model ◽

Dynamics Simulations

The deamination process of isoxanthopterin catalyzed by isoxanthopterin deaminase was determined using the combined QM(PM3)/MM molecular dynamics simulations. In this paper, the updated PM3 parameters were employed for zinc ions and the initial model was built up based on the crystal structure. Proton transfer and following steps have been investigated in two paths: Asp336 and His285 serve as the proton shuttle, respectively. Our simulations showed that His285 is more effective than Aap336 in proton transfer for deamination of isoxanthopterin. As hydrogen bonds between the substrate and surrounding residues play a key role in nucleophilic attack, we suggested mutating Thr195 to glutamic acid, which could enhance the hydrogen bonds and help isoxanthopterin get close to the active site. The simulations which change the substrate to pterin 6-carboxylate also performed for comparison. Our results provide reference for understanding of the mechanism of deaminase and for enhancing the deamination rate of isoxanthopterin deaminase.

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Exploiting correlated molecular-dynamics networks to counteract enzyme activity–stability trade-off

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1812204115 ◽

2018 ◽

Vol 115 (52) ◽

pp. E12192-E12200 ◽

Cited By ~ 13

Author(s):

Haoran Yu ◽

Paul A. Dalby

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Active Site ◽

Dynamic Networks ◽

Aromatic Aldehydes ◽

Active Site Residues ◽

Trade Off ◽

Highly Correlated ◽

Dynamics Simulations ◽

The Impact

The directed evolution of enzymes for improved activity or substrate specificity commonly leads to a trade-off in stability. We have identified an activity–stability trade-off and a loss in unfolding cooperativity for a variant (3M) of Escherichia coli transketolase (TK) engineered to accept aromatic substrates. Molecular dynamics simulations of 3M revealed increased flexibility in several interconnected active-site regions that also form part of the dimer interface. Mutating the newly flexible active-site residues to regain stability risked losing the new activity. We hypothesized that stabilizing mutations could be targeted to residues outside of the active site, whose dynamics were correlated with the newly flexible active-site residues. We previously stabilized WT TK by targeting mutations to highly flexible regions. These regions were much less flexible in 3M and would not have been selected a priori as targets using the same strategy based on flexibility alone. However, their dynamics were highly correlated with the newly flexible active-site regions of 3M. Introducing the previous mutations into 3M reestablished the WT level of stability and unfolding cooperativity, giving a 10.8-fold improved half-life at 55 °C, and increased midpoint and aggregation onset temperatures by 3 °C and 4.3 °C, respectively. Even the activity toward aromatic aldehydes increased up to threefold. Molecular dynamics simulations confirmed that the mutations rigidified the active-site via the correlated network. This work provides insights into the impact of rigidifying mutations within highly correlated dynamic networks that could also be useful for developing improved computational protein engineering strategies.

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UNDERSTANDING DIFFERENT LCST LEVELS OF POLY(N-ALKYLACRYLAMIDE)S BY MOLECULAR DYNAMICS SIMULATIONS AND QUANTUM MECHANICS CALCULATIONS

Journal of Theoretical and Computational Chemistry ◽

10.1142/s0219633611006505 ◽

2011 ◽

Vol 10 (03) ◽

pp. 359-370 ◽

Cited By ~ 7

Author(s):

JUAN PANG ◽

HU YANG ◽

JING MA ◽

RONGSHI CHENG

Keyword(s):

Molecular Dynamics ◽

Quantum Mechanics ◽

Aqueous Solutions ◽

Molecular Dynamics Simulations ◽

Steric Effect ◽

Conformational Transition ◽

Methyl Group ◽

Branched Chain ◽

Dynamics Simulations ◽

Different Levels

Poly(N-alkylacrylamide) is a group of thermo-sensitive polymers that include poly (N-isopropylacrylamide), poly(N-n-propylacrylamide), poly(N-isopropylmethacryl-amide), and so on. The polymers exhibit different levels of lower critical solution temperatures (LCST) in aqueous solutions. In this article, their monomers and oligomers with 10 repeating units are selected, respectively, to demonstrate the cause of different LCST levels of the polymers in aqueous solutions using molecular dynamics simulations and quantum mechanics calculations. The monomers have functional groups of different steric volume that greatly affect the conformational transition of chains and LCST levels of the polymers. A branched chain of N-propyl group in N-isopropylacrylamide and an additional methyl group at α-carbon in N-isopropylmethacrylamide both increase the steric effect, making it more difficult for monomers to draw closer and resulting in higher LCST levels of the polymers. In addition, the simulated results from their corresponding oligomers exhibit the similar trend to those from the monomers.

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Chain dynamics and conformational transition in cis-polyisoprene: Comparison between melt and subglass state by molecular dynamics simulations

The Journal of Chemical Physics ◽

10.1063/1.1288023 ◽

2000 ◽

Vol 113 (10) ◽

pp. 4433-4443 ◽

Cited By ~ 15

Author(s):

Mitsuhiro Fukuda ◽

Hiroaki Kikuchi

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Conformational Transition ◽

Chain Dynamics ◽

Dynamics Simulations ◽

In Cis

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Effects of temperatures on pressure-induced structural changes in amorphous Si: A molecular-dynamics study

10.29007/6kp3 ◽

2020 ◽

Author(s):

Renji Mukuno ◽

Manabu Ishimaru

Keyword(s):

Molecular Dynamics ◽

Phase Transformation ◽

Amorphous Phase ◽

Structural Changes ◽

Interatomic Potential ◽

Activation Barrier ◽

Distribution Functions ◽

High Density ◽

Angle Distribution ◽

Dynamics Simulations

The structural changes of amorphous silicon (a-Si) under compressive pressure were examined by molecular-dynamics simulations using the Tersoff interatomic potential. a-Si prepared by melt-quenching methods was pressurized up to 30 GPa under different temperatures (300K and 500K). The density of a-Si increased from 2.26 to 3.24 g/cm3 with pressure, suggesting the occurrence of the low-density to high-density amorphous phase transformation. This phase transformation occurred at the lower pressure with increasing the temperature because the activation barrier for amorphous-to-amorphous phase transformation could be exceeded by thermal energy. The coordination number increased with pressure and time, and it was saturated at different values depending on the pressure. This suggested the existence of different metastable atomic configurations in a-Si. Atomic pair-distribution functions and bond-angle distribution functions suggested that the short-range ordered structure of high-density a-Si is similar to the structure of the high-pressure phase of crystalline Si (β-tin and Imma structures).

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