Enhanced enzymatic activity exerted by a packed assembly of a single type of enzyme

Dinh, Huyen; Nakata, Eiji; Mutsuda-Zapater, Kaori; Saimura, Masayuki; Kinoshita, Masahiro; Mutoh, T.

doi:10.1039/d0sc03498c

Cited by 15 publications

(8 citation statements)

References 62 publications

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“…The characteristics of HPO and HPC are quite useful for the applications of the dynamic DNA scaffold as the nanocarrier for enzymes or other macromolecules to maintain the catalytic activity of assembled enzyme during the shape transformation, in which the catalytic activity of the enzyme on HPO is higher than its free form ( Lin et al, 2021 ). Moreover, the enhancement of catalytic activity would be further tuned by assembling the same type of enzymes in the packed state ( Dinh et al, 2020 ).…”

Section: Resultsmentioning

confidence: 99%

Dynamic Shape Transformation of a DNA Scaffold Applied for an Enzyme Nanocarrier

et al. 2021

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Structural programmability and accurate addressability of DNA nanostructures are ideal characteristics for the platform of arranging enzymes with the nanoscale precision. In this study, a three-dimensional DNA scaffold was designed to enable a dynamic shape transition from an open plate-like structure to its closed state of a hexagonal prism structure. The two domains in the open state were folded together to transform into the closed state by hybridization of complementary short DNA closing keys at both of the facing edges in over 90% yield. The shape transformation of the DNA scaffold was extensively studied by means of the fluorescence energy transfer measurement, atomic force microscope images, and agarose gel electrophoretic analyses. A dimeric enzyme xylitol dehydrogenase was assembled on the DNA scaffold in its open state in a high-loading yield. The enzyme loaded on the scaffold was subsequently transformed to its closed state by the addition of short DNA closing keys. The enzyme encapsulated in the closed state displayed comparable activity to that in the open state, ensuring that the catalytic activity of the enzyme was well maintained in the DNA nanocarrier. The nanocarrier with efficient encapsulation ability is potentially applicable for drug delivery, biosensing, biocatalytic, and diagnostic tools.

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Section: Resultsmentioning

confidence: 99%

Dynamic Shape Transformation of a DNA Scaffold Applied for an Enzyme Nanocarrier

et al. 2021

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“…Generally, hydrophobic interaction is one of the most important driving forces for protein oligomerization, which benefits their temperature stability ( Vieille and Zeikus, 2001 ). In addition, it has been demonstrated that the enzymes in the packed state showed higher catalytic activity than those in the dispersed state due to increases in local substrate or cofactor concentration within the domain between enzyme surfaces, resulting from water entropic force ( Dinh et al, 2020 ). Therefore, the hydrophobic interaction-mediated oligomerization or aggregation could play an essential role in the stabilization and activity enhancement of EAK16-E3C/E55F/D213T.…”

Section: Discussionmentioning

confidence: 99%

Enhanced salt-tolerance of Bacillus subtilis glutaminase by fusing self-assembling amphipathic peptides at its N-terminus

Liu

Rao

Chen

et al. 2022

Front. Bioeng. Biotechnol.

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Glutaminase (EC 3.5.1.2) can catalyze the deamidation of glutamine, which has been used to improve umami taste in oriental fermented foods. However, a high salt concentration is still a fundamental challenge for glutaminase application, especially in soy sauce production. To improve the salt tolerance of glutaminase, the self-assembling amphiphilic peptides EAK16 and ELK16 were fused to the N-terminus of a mutant (E3C/E55F/D213T) derived from Bacillus subtilis glutaminase, yielding the fusion enzymes EAK16-E3C/E55F/D213T and ELK16-E3C/E55F/D213T, respectively. As ELK16-E3C/E55F/D213T was expressed as insoluble active inclusion bodies, only the purified EAK16-E3C/E55F/D213T was subjected to further analyses. After the incubation with 18% (w/v) NaCl for 200 min, the residual activities of EAK16-E3C/E55F/D213T in a NaCl-free solution reached 43.6%, while E3C/E55F/D213T was completely inactivated. When the enzyme reaction was conducted in the presence of 20% NaCl, the relative activity of EAK16-E3C/E55F/D213T was 0.47-fold higher than that of E3C/E55F/D213T. As protein surface hydrophobicity and protein particle size analysis suggested, oligomerization may play an important role in the salt-tolerance enhancement of the fusions. Furthermore, EAK16-E3C/E55F/D213T achieved a 0.88-fold increase in the titer of glutamic acid in a model system of soy sauce fermentation compared to E3C/E55F/D213T. Therefore, the fusion with self-assembling amphiphilic peptides is an efficient strategy to improve the salt-tolerance of glutaminase.

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“…Highly dense packing of HRP on the surface of nano-assembly might result in partial thermal denaturation to decrease the activity. However, Morii and coworkers reported a surprising enhancement of activity of dense-packed assembly of carbonic anhydrase on DNA origami scaffolds [47]. They proposed that the entropic force of water increases local substrate or cofactor concentration within the domain confined between enzyme surfaces, thus accelerating the reaction.…”

Section: Discussionmentioning

confidence: 99%

Horseradish Peroxidase-Decorated Artificial Viral Capsid Constructed from β-Annulus Peptide via Interaction between His-Tag and Ni-NTA

et al. 2020

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Artificial construction of spherical protein assemblies has attracted considerable attention due to its potential use in nanocontainers, nanocarriers, and nanoreactors. In this work, we demonstrate a novel strategy to construct peptide nanocapsules (artificial viral capsids) decorated with enzymes via interactions between His-tag and Ni-NTA. A β-annulus peptide derived from the tomato bushy stunt virus was modified with Ni-NTA at the C-terminus, which is directed toward the exterior surface of the artificial viral capsid. The β-annulus peptide bearing Ni-NTA at the C-terminus self-assembled into capsids of about 50 nm in diameter. The Ni-NTA-displayed capsids were complexed with recombinant horseradish peroxidase (HRP) with a C-terminal His-tag which was expressed in Escherichia coli. The β-annulus peptide-HRP complex formed spherical assemblies whose sizes were 30–90 nm, with the ζ-potential revealing that the HRP was decorated on the outer surface of the capsid.

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Enhanced enzymatic activity exerted by a packed assembly of a single type of enzyme

Abstract: In contrast to the dilute conditions employed for in vitro biochemical studies, enzymes are spatially organized at high density in cellular micro-compartments. In spite of being crucial for cellular functions,...

Cited by 15 publications

References 62 publications

Dynamic Shape Transformation of a DNA Scaffold Applied for an Enzyme Nanocarrier

Dynamic Shape Transformation of a DNA Scaffold Applied for an Enzyme Nanocarrier

Enhanced salt-tolerance of Bacillus subtilis glutaminase by fusing self-assembling amphipathic peptides at its N-terminus

Horseradish Peroxidase-Decorated Artificial Viral Capsid Constructed from β-Annulus Peptide via Interaction between His-Tag and Ni-NTA

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