The multicopper oxidase enzyme laccase holds great potential to be used for biological lignin valorization alongside a biocompatible ionic liquid (IL). However, the IL concentrations required for biomass pretreatment severely inhibit laccase activity. Due to their ability to function in extreme conditions, many thermophilic enzymes have found use in industrial applications. The thermophilic fungal laccase from Myceliophthora thermophila was found to retain high levels of activity in the IL [C 2 C 1 Im][EtSO 4 ], making it a desirable biocatalyst to be used for lignin valorization. In contrast to [C 2 C 1 Im][EtSO 4 ], the biocompatibility of [C 2 C 1 Im][OAC] with the laccase was markedly lower. Severe inhibition of laccase activity was observed in 15% [C 2 C 1 Im][OAc]. In this study, the enzyme surface charges were modified via acetylation, succinylation, cationization, or neutralization. However, these modifications did not show significant improvement in laccase activity or stability in [C 2 C 1 Im][OAc]. Docking simulations show that the IL docks close to the T1 catalytic copper, likely interfering with substrate binding. Although additional docking locations for [OAc] - are observed after making enzyme modifications, it does not appear that these locations play a role in the inhibition of enzyme activity. The results of this study could guide future enzyme engineering efforts by showing that the inhibition mechanism of [C 2 C 1 Im][OAc] toward M. thermophila laccase is likely not dependent upon the IL interacting with the enzyme surface.
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Effects of Ionic Liquids on Laccase from Trametes versicolor
Interactions between ionic liquids and biomolecules are of great interest due to the intrinsic properties of ionic liquids and the flexibility allowed by mixing and matching cations and anions to create unique ionic liquids. A number of ionic liquid–biomolecule studies have focused on interactions with proteins, including industrially relevant enzymes. One of these, laccase from Trametes versicolor, is a naturally derived enzyme used in the breakdown of phenolic compounds in a wide variety of industries, especially useful in breakdown of lignocellulosic materials. Here, a combination of experiments and molecular dynamics (MD) simulations was used to investigate the interactions of ionic liquids with laccase. Enzyme kinetics assays indicated that ionic liquids composed of tetramethylguanidine (TMG) and either serine or threonine caused significant reduction in enzymatic activity, while kinetics was not impacted by TMG-Asp or TMG-Glu ionic liquids. Similarly, intrinsic fluorescence of laccase in the presence of TMG-Ser and TMG-Thr exhibited a shift in spectral properties consistent with structural destabilization, but again TMG-Asp and TMG-Glu had no impact. MD simulations of laccase and ABTS with/without TMG-Ser ionic liquid provided insight into the deactivation mechanism of laccase. The simulations indicated that TMG-Ser disrupts laccase’s electron transfer mechanism.
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- Award ID(s):
- 1904797
- NSF-PAR ID:
- 10332942
- Date Published:
- Journal Name:
- Biophysica
- Volume:
- 1
- Issue:
- 4
- ISSN:
- 2673-4125
- Page Range / eLocation ID:
- 429 to 444
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Abstract Ionic liquid (IL)‐containing polymers garner attention for electrochemical applications. This article overviews recent experimental and theoretical studies of polymer electrolytes that would be likely to cultivate new theoretical and computational frameworks for IL‐containing polymers. The first two sections outline the uniqueness of ILs that differentiates them from inorganic salts in polymers and explore deviation from the concept of the metaphor “room‐temperature molten salt.” Such distinct properties include (1) large intrinsic dipole moment and electronic polarizability, (2) hydrogen bonding, (3) π‐interactions, (4) a broad distribution of charges over the entire ion, and (5) the anisotropy of the ions. Moreover, the complexity of these properties substantially increases when the ions are polymerized. Indeed, their exceptional features would overcome the hurdle due to a trade‐off between ionic conductivity and mechanical robustness in inorganic salt‐doped polymers. Given these facts, the rest of the article focuses on emerging trends in the study of the dielectric response, phase separation, ion conductivity, and mechanical robustness of the polymer electrolytes, highlighting outstanding observations in experiments that may inspire existing theory and simulation. Our discussion also includes improving computational complexity for IL‐containing polymers. To this end, recent machine learning studies that consider ILs and polymer liquids are presented.
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Abstract DNA glycosylase MutY plays a critical role in suppression of mutations resulted from oxidative damage, as highlighted by cancer-association of the human enzyme. MutY requires a highly conserved catalytic Asp residue for excision of adenines misinserted opposite 8-oxo-7,8-dihydroguanine (OG). A nearby Asn residue hydrogen bonds to the catalytic Asp in structures of MutY and its mutation to Ser is an inherited variant in human MUTYH associated with colorectal cancer. We captured structural snapshots of N146S Geobacillus stearothermophilus MutY bound to DNA containing a substrate, a transition state analog and enzyme-catalyzed abasic site products to provide insight into the base excision mechanism of MutY and the role of Asn. Surprisingly, despite the ability of N146S to excise adenine and purine (P) in vitro, albeit at slow rates, N146S-OG:P complex showed a calcium coordinated to the purine base altering its conformation to inhibit hydrolysis. We obtained crystal structures of N146S Gs MutY bound to its abasic site product by removing the calcium from crystals of N146S-OG:P complex to initiate catalysis in crystallo or by crystallization in the absence of calcium. The product structures of N146S feature enzyme-generated β-anomer abasic sites that support a retaining mechanism for MutY-catalyzed base excision.
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ABSTRACT The use of ionic liquids (ILs) as media in radical polymerizations has demonstrated the ability of these unique solvents to improve both reaction kinetics and polymer product properties. However, the bulk of these studies have examined the polymerization behavior of common organic monomers (e.g., methyl methacrylate, styrene) dissolved in conventional ILs. There is increasing interest in polymerized ILs (poly(ILs)), which are ionomers produced from the direct polymerization of styrene‐, vinyl‐, and acrylate‐functionalized ILs. Here, the photopolymerization kinetics of IL monomers are investigated for systems in which styrene or vinyl functionalities are pendant from the imidazolium cation. Styrene‐functionalized IL monomers typically polymerized rapidly (full conversion ≤1 min) in both neat compositions or when diluted with a nonpolymerizable IL, [C2mim][Tf2N]. However, monomer conversion in vinyl‐functionalized IL monomers is much more dependent on the nature of the nonpolymerizable group. ATR‐FTIR analysis and molecular simulations of these monomers and monomer mixtures identified the presence of multiple intermolecular interactions (e.g., π–π stacking, IL aggregation) that contribute to the polymerization behaviors of these systems. © 2018 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem.
2018 ,56 , 2364–2375
- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3390/biophysica1040031
