Apoenzyme Reconstitution as a Chemical Tool for Structural Enzymology and Biotechnology

Fruk, Ljiljana; Kuo, Chih‐Yu; Torres, Eduardo; Niemeyer, Christof M.

doi:10.1002/anie.200803098

Cited by 117 publications

(78 citation statements)

References 226 publications

(243 reference statements)

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“…The electrical contacting of redox proteins with electrodes attracted substantial research efforts directed to the development of amperometric biosensors [26][27][28][29] or biofuel cell elements [30][31][32][33][34][35] . Different methods to integrate redox proteins with electrodes in electrically contacted configurations were developed, including the modification of the proteins with relay units 36,37 , the reconstitution of the proteins on relay-cofactor units [38][39][40] and the immobilization of the proteins in redox-active polymer matrices 41 . Related principles were implemented to electrically wire PSII with electrode surfaces.…”

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confidence: 99%

Integrated photosystem II-based photo-bioelectrochemical cells

et al. 2012

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Photosynthesis is a sustainable process that converts light energy into chemical energy. substantial research efforts are directed towards the application of the photosynthetic reaction centres, photosystems I and II, as active components for the light-induced generation of electrical power or fuel products. nonetheless, no integrated photo-bioelectrochemical device that produces electrical power, upon irradiation of an aqueous solution that includes two inter-connected electrodes is known. Here we report the assembly of photobiofuel cells that generate electricity upon irradiation of biomaterial-functionalized electrodes in aqueous solutions. The cells are composed of electrically contacted photosystem II-functionalized photoanodes and an electrically wired bilirubin oxidase/carbon nanotubes-modified cathode. Illumination of the photoanodes yields the oxidation of water to o 2 and the transfer of electrons through the external circuit to the cathode, where o 2 is re-reduced to water.

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confidence: 99%

Integrated photosystem II-based photo-bioelectrochemical cells

et al. 2012

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“…alpha-hemolysin | single molecule | stochastic sensing | structure | unnatural amino acid M any research groups have used protein engineering to obtain enzymes and antibodies with new properties suited for specific tasks (1)(2)(3)(4)(5)(6). Fewer groups have taken on the difficult problem of engineering membrane proteins (7).…”

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confidence: 99%

Molecular bases of cyclodextrin adapter interactions with engineered protein nanopores

Banerjee

Mikhailova

Cheley

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

108

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Engineered protein pores have several potential applications in biotechnology: as sensor elements in stochastic detection and ultrarapid DNA sequencing, as nanoreactors to observe singlemolecule chemistry, and in the construction of nano-and microdevices. One important class of pores contains molecular adapters, which provide internal binding sites for small molecules. Mutants of the α-hemolysin (αHL) pore that bind the adapter β-cyclodextrin (βCD) ∼10 4 times more tightly than the wild type have been obtained. We now use single-channel electrical recording, protein engineering including unnatural amino acid mutagenesis, and highresolution x-ray crystallography to provide definitive structural information on these engineered protein nanopores in unparalleled detail.alpha-hemolysin | single molecule | stochastic sensing | structure | unnatural amino acid M any research groups have used protein engineering to obtain enzymes and antibodies with new properties suited for specific tasks (1-6). Fewer groups have taken on the difficult problem of engineering membrane proteins (7). We have engineered the α-hemolysin protein pore, mindful of several potential applications in biotechnology, including its ability to act as a detector in stochastic sensing (8) and ultrarapid DNA sequencing (9), to serve as a nanoreactor for the observation of singlemolecule chemistry (10) and to act as a component for the construction of nano-and microdevices (11).An important breakthrough in this area, which enabled the stochastic sensing of organic molecules including the detection of DNA bases in the form of nucleoside monophosphates (12, 13), was the discovery of internal molecular adapters, a form of noncovalent protein modification (14). Most useful have been cyclodextrin (CD) adapters, which have until now been used in the absence of detailed structural information about how they work. The present paper is a definitive investigation, which provides such information through the application of a wide variety of technical approaches: single-channel electrical recording, protein engineering including unnatural amino acid mutagenesis, and x-ray crystallography. The studies employing mutagenesis show that the striking interactions seen in the crystal structures also occur in individual pores in lipid bilayers.We reveal that the tight-binding αHL mutants (15) M113N 7 and M113F 7 bind βCD in different orientations within the heptameric pore. In the case of M113N 7 , the top (primary hydroxyls) of the CD ring faces the trans entrance of the pore. In the case of M113F 7 , the bottom (secondary hydroxyls) of the CD ring faces the trans entrance, while the top of the ring is bonded to the pore through remarkable CH-π interactions. Another tight-binding mutant, M113V 7 , can bind the CD in both orientations. These results illustrate the exquisite level of engineering that can be achieved with protein nanopores, which is, for example, far beyond what is possible with solid-state pores. The work also provides information valuable for the design of ...

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“…6 In such way novel biomaterials were designed by use of DNA- 19 and polymer modified heme groups. 20 In many cases, enhanced catalytic performance was enabled by modification of the heme propionate groups, 21 exchange of iron centre 22 or by appending electro- 23 and photoactive 8,24 groups to these side chains.…”

Section: Resultsmentioning

confidence: 99%

“…6 As a result, two products are formed, namely heme derivatives where one or both propionic groups are modified with a DNA strand (2 and 3 respectively in Figure 1). The CPG was then washed and reacted with an access of amino-bpy 4, synthesized according to a previously reported procedure, to yield heme derivative 5 ( Figure 1).…”

Section: Resultsmentioning

confidence: 99%

“…This is achieved either by modification of theprotein shell through introduction of unnatural amino acids 5 or by replacing metal containing prosthetic groups with artificial ones. 6 Since tailoring of the protein shell often requires lengthy process of site selective mutations and subsequent enzyme expression, this approach is often of limited because of the low amounts of the biocatalyst that can be produced. 5 On the other hand, the modification of small organic prosthetic group offers ease of preparation and a range of different functions, which can be introduced.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Bimetallic Copper-Heme-Protein-DNA Hybrid Catalyst for Diels Alder Reaction

Kuo¹,

Niemeyer²,

Fruk³

2011

Croat. Chem. Acta

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Abstract.A bimetallic heme-DNA cofactor, containing an iron and a copper center, was synthesized for the design of novel hybrid catalysts for stereoselective synthesis. The cofactor was used for the reconstitution of apo-myoglobin. Both the cofactor alone and its myoglobin adduct were used to catalyze a model Diels Alder reaction. Stereoselectivity of this conversion was analyzed by chiral HPLC. Reactions carried out in the presence of myoglobin-heme-Cu-DNA catalyst showed greater product conversion and stereoselectivity than those carried out with the heme-Cu-DNA cofactor. This observation suggested that the protein shell plays a significant role in the catalytic conversion.(doi: 10.5562/cca1828)

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Apoenzyme Reconstitution as a Chemical Tool for Structural Enzymology and Biotechnology

Cited by 117 publications

References 226 publications

Integrated photosystem II-based photo-bioelectrochemical cells

Integrated photosystem II-based photo-bioelectrochemical cells

Molecular bases of cyclodextrin adapter interactions with engineered protein nanopores

Bimetallic Copper-Heme-Protein-DNA Hybrid Catalyst for Diels Alder Reaction

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