Molecular mechanism of enzyme tolerance against organic solvents: Insights from molecular dynamics simulation

Mohtashami, Mahnaz; Fooladi, Jamshid; Haddad-Mashadrizeh, Aliakbar; Housaindokht, Mohammad Reza; Monhemi, Hassan

doi:10.1016/j.ijbiomac.2018.10.172

Cited by 56 publications

(36 citation statements)

References 48 publications

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“…Methanol is instead a hydrogen-bonding donor, which can form H-bonds with the backbone CO-group [25]. At the same time, both DMSO and methanol do not significantly affect side-chain hydrogen bonds and salt bridges [24,25,29]. The analysis of hydrogen bonds in the functional dimers of TaTT homologs showed that TaTT is distinguished by the excess of surface hydrogen bonds, by hydrogen bonds formed by side chains of charged residues (salt bridges) and by the number of hydrogen bonds per one amino acid residue ( Figure 5, Table A1).…”

Section: Scheme 1 Transamination Reaction Between L-leu and α-Ketoglmentioning

confidence: 99%

“…The solvent-induced structural changes can alter the conformational flexibility of the enzyme and adjust the balance between rigidity and flexibility at suboptimal temperatures, i.e., lower than 75 • C. To interpret the effects of the organic solvents on the thermostability and thermophilicity of TaTT, we focused on the analysis of hydrogen bonding in TaTT, considering hydrogen-bonding network as a crucial structural factor of the stability of enzymes in harsh conditions. According to the recent studies, the penetration of solvent molecules into the hydration shell of an enzyme molecule disturbs the surface hydrogen bonds because of the amphiphilic nature of the cosolvents (dipole strength is 3.96 D for DMSO, 1.70 D for methanol, and 1.85 D for water) [24,26,29]. Both DMSO and methanol can interact with the backbone atoms and outcompete water molecules to form surface hydrogen bonds [24][25][26]33,39].…”

Section: Scheme 1 Transamination Reaction Between L-leu and α-Ketoglmentioning

confidence: 99%

“…According to the recent studies, the penetration of solvent molecules into the hydration shell of an enzyme molecule disturbs the surface hydrogen bonds because of the amphiphilic nature of the cosolvents (dipole strength is 3.96 D for DMSO, 1.70 D for methanol, and 1.85 D for water) [24,26,29]. Both DMSO and methanol can interact with the backbone atoms and outcompete water molecules to form surface hydrogen bonds [24][25][26]33,39]. Present in the hydration shell, DMSO can interact with the exposed backbone NH-groups via its oxygen atoms [29].…”

Section: Scheme 1 Transamination Reaction Between L-leu and α-Ketoglmentioning

confidence: 99%

“…In particular, it was observed that solvent molecules replaced water molecules on the surface and even in the active site of enzymes [17][18][19][20][21]. It is considered that water-miscible organic solvent-water mixtures destabilize the protein globule: solvent molecules penetrate the hydration shell of the enzyme molecules and strip water from the surface, thus solvating hydrophobic patches and disrupting the surface hydrogen bonds due to the interaction with backbone atoms [7,[22][23][24][25][26]. A water-miscible organic solvent can interfere with the activity of an enzyme by substituting catalytic water molecules and breaking conserved hydrogen bonds in the active site [27,28].…”

Section: Introductionmentioning

confidence: 99%

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Counterbalance of Stability and Activity Observed for Thermostable Transaminase from Thermobaculum terrenum in the Presence of Organic Solvents

et al. 2020

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Pyridoxal-5’-phosphate-dependent transaminases catalyze stereoselective amination of organic compounds and are highly important for industrial applications. Catalysis by transaminases often requires organic solvents to increase the solubility of reactants. However, natural transaminases are prone to inactivation in the presence of water-miscible organic solvents. Here, we present the solvent tolerant thermostable transaminase from Thermobaculum terrenum (TaTT) that catalyzes transamination between L-leucine and alpha-ketoglutarate with an optimum at 75 °C and increases the activity ~1.8-fold upon addition of 15% dimethyl sulfoxide or 15% methanol at high but suboptimal temperature, 50 °C. The enhancement of the activity correlates with a decrease in the thermal denaturation midpoint temperature. The blue-shift of tryptophan fluorescence suggested that solvent molecules penetrate the hydration shell of the enzyme. Analysis of hydrogen bonds in the TaTT dimer revealed a high number of salt bridges and surface hydrogen bonds formed by backbone atoms. The latter are sensitive to the presence of organic solvents; they rearrange, conferring the relaxation of some constraints inherent to a thermostable enzyme at low temperatures. Our data support the idea that the counterbalance of stability and activity is crucial for the catalysis under given conditions; the obtained results may be useful for fine-tuning biocatalyst efficiency.

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Section: Scheme 1 Transamination Reaction Between L-leu and α-Ketoglmentioning

confidence: 99%

Section: Scheme 1 Transamination Reaction Between L-leu and α-Ketoglmentioning

confidence: 99%

Section: Scheme 1 Transamination Reaction Between L-leu and α-Ketoglmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Counterbalance of Stability and Activity Observed for Thermostable Transaminase from Thermobaculum terrenum in the Presence of Organic Solvents

et al. 2020

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“…In addition, the efficacy of lysozyme is limited to Gram-positive bacteria due to the property of lysozyme targeting the β-1,4 glycosidic linkages between N-acetylglucosamine and N-acetylmuraminic acid of peptidoglycan. It has been reported that modifying lysozyme can change this limitation, including chemical, physical, and biological changes [15,16]. Among all the strategies, the immobilization of enzymes has attracted considerable interest [17].…”

Section: Introductionmentioning

confidence: 99%

Enhancing the Thermo-Stability and Anti-Bacterium Activity of Lysozyme by Immobilization on Chitosan Nanoparticles

Wang

Jin

et al. 2020

IJMS

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The recent emergence of antibiotic-resistant bacteria requires the development of new antibiotics or new agents capable of enhancing antibiotic activity. Lysozyme degrades bacterial cell wall without involving antibiotic resistance and has become a new antibacterial strategy. However, direct use of native, active proteins in clinical settings is not practical as it is fragile under various conditions. In this study, lysozyme was integrated into chitosan nanoparticles (CS-NPs) by the ionic gelation technique to obtain lysozyme immobilized chitosan nanoparticles (Lys-CS-NPs) and then characterized by dynamic light scattering (DLS) and transmission electron microscopy (TEM), which showed a small particle size (243.1 ± 2.1 nm) and positive zeta potential (22.8 ± 0.2 mV). The immobilization significantly enhanced the thermal stability and reusability of lysozyme. In addition, compared with free lysozyme, Lys-CS-NPs exhibited superb antibacterial properties according to the results of killing kinetics in vitro and measurement of the minimum inhibitory concentration (MIC) of CS-NPs and Lys-CS-NPs against Pseudomonas aeruginosa (P. aeruginosa), Klebsiella pneumoniae (K. pneumoniae), Escherichia coli (E. coli), and Staphylococcus aureus (S. aureus). These results suggest that the integration of lysozyme into CS-NPs will create opportunities for the further potential applications of lysozyme as an anti-bacterium agent.

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Semirational engineering of an aldo–keto reductase KmAKR for overcoming trade‐offs between catalytic activity and thermostability

Xie

Qiu

et al. 2021

Biotech & Bioengineering

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Enzyme engineering usually generates trade-offs between activity, stability, and selectivity. Herein, we report semirational engineering of an aldo-keto reductase (AKR) KmAKR for simultaneously enhancing its thermostability and catalytic activity.Previously, we constructed KmAKR M9 (W297H/Y296W/K29H/Y28A/T63M/A30P/ T302S/N109K/S196C), which showed outstanding activity towards t-butyl 6-chloro-(3R,5S)-dihydroxyhexanoate ((3R,5S)-CDHH), and t-butyl 6-cyano-(3R,5R)dihydroxyhexanoate, the key chiral building blocks of rosuvastatin and atorvastatin.Under the guidance of computer-aided design including consensus residues analysis and molecular dynamics (MD) simulations, K164, S182, S232, and Q266 were dug out for their thermostability conferring roles, generating the "best" mutant KmAKR M13 (W297H/Y296W/K29H/Y28A/T63M/A30P/T302S/N109K/S196C/ K164E/S232A/S182H/Q266D). The T m and T 50 15 values of KmAKR M13 were 10.4 and 6.1°C higher than that of KmAKR M9 , respectively. Moreover, it displayed a significantly elevated organic solvent tolerance over KmAKR M9 . Structural analysis indicated that stabilization of the α-helixes mainly contributed to thermostability enhancement. Under the optimized conditions, KmAKR M13 completely asymmetrically reduced 400 g/l t-butyl 6-chloro-(5S)-hydroxy-3-oxohexanoate ((5S)-CHOH) in 8.0 h at a high substrate to catalyst ratio (S/C) of 106.7 g/g, giving diastereomerically pure (3R,5S)-CDHH (>99.5% d.e. P ) with a space-time yield (STY) of 449.2 g/l•d.

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Molecular mechanism of enzyme tolerance against organic solvents: Insights from molecular dynamics simulation

Cited by 56 publications

References 48 publications

Counterbalance of Stability and Activity Observed for Thermostable Transaminase from Thermobaculum terrenum in the Presence of Organic Solvents

Counterbalance of Stability and Activity Observed for Thermostable Transaminase from Thermobaculum terrenum in the Presence of Organic Solvents

Enhancing the Thermo-Stability and Anti-Bacterium Activity of Lysozyme by Immobilization on Chitosan Nanoparticles

Semirational engineering of an aldo–keto reductase KmAKR for overcoming trade‐offs between catalytic activity and thermostability

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