A Dual Anchoring Strategy for the Localization and Activation of Artificial Metalloenzymes Based on the Biotin–Streptavidin Technology

Zimbron, Jeremy M.; Heinisch, Tillmann; Schmid, Maurus; Hamels, Didier; Nogueira, Elisa S.; Schirmer, Tilman; Ward, Thomas R.

doi:10.1021/ja309974s

Cited by 92 publications

(74 citation statements)

References 32 publications

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“…[12,19,28,29] The dimeric precursor was reacted in situ with a selection of commercially available bidentate ligands (Scheme 2). To identify suitable coordination conditions, the chiral ligand l-ProNH 2 was complexed in situ with the dinuclear catalyst precursors [MCp* biotin Cl 2 ] 2 (M = Ir, Rh) at different pH values within the buffer range of 3-(N-morpholino)propanesulfonic acid (MOPS) or 2-(N-morpholino)ethanesulfonic acid.…”

Section: Resultsmentioning

confidence: 99%

“…[1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] The resulting hybrid catalyst can be optimized by using either genetic or chemical methods. [1][2][3][4][5][6][7][8][18][19][20][21] In the context of artificial metalloenzymes based on the biotin-streptavidin technology, we and others have relied on synthesizing various biotinylated ligands to provide chemical diversity. [1,4,15,20,21] To generate larger artificial cofactor libraries quickly, we reasoned that separating the necessary biotin anchor moiety and variable ligand elements would enable us to screen commercially available ligands in the presence of streptavidin (Sav; Scheme 1).…”

Section: Introductionmentioning

confidence: 99%

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Expanding the Chemical Diversity in Artificial Imine Reductases Based on the Biotin–Streptavidin Technology

Quinto¹,

Schwizer²,

Zimbron³

et al. 2014

ChemCatChem

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We report on the optimization of an artificial imine reductase based on the biotin‐streptavidin technology. With the aim of rapidly generating chemical diversity, a novel strategy for the formation and evaluation of biotinylated complexes is disclosed. Tethering the biotin‐anchor to the Cp* moiety leaves three free coordination sites on a d6 metal for the introduction of chemical diversity by coordination of a variety of ligands. To test the concept, 34 bidentate ligands were screened and a selection of the 6 best was tested in the presence of 21 streptavidin (Sav) isoforms for the asymmetric imine reduction by the resulting three legged piano stool complexes. Enantiopure α‐amino amides were identified as promising bidentate ligands: up to 63 % ee and 190 turnovers were obtained in the formation of 1‐phenyl‐1,2,3,4‐tetrahydroisoquinoline with [IrCp*biotin(L‐ThrNH2)Cl]⊂SavWT as a catalyst.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Expanding the Chemical Diversity in Artificial Imine Reductases Based on the Biotin–Streptavidin Technology

Quinto¹,

Schwizer²,

Zimbron³

et al. 2014

ChemCatChem

Self Cite

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“…[53] When histidine was introduced at S112 (i.e.S112H), the conversion of the reaction increases 10-fold over the wild-type complex to provide (S)-salsolidine (29)i n quantitative yield and 55 % ee.I nterestingly,t he enantioselectivity is inverted when histidine was introduced at position K121 (K121H), with (R)-salsolidine (29)g enerated in quantitative yield and 79 % ee (Figure 15). Evaluation of the crystal structures of the two variants support the metal adopting very different orientations within the chiral biotinbinding vestibule depending on the position of the histidine residue,potentially accounting for the opposite enantioselectivities.Although, in general, metalloenzymes rely on genetic modification of the second coordination sphere,t his study suggests that genetic optimization of the first coordination sphere can be used to enhance both the reactivity and the selectivity of artificial metalloenzymes.…”

Section: Angewandte Chemiementioning

confidence: 99%

Genetic Optimization of Metalloenzymes: Enhancing Enzymes for Non‐Natural Reactions

2016

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Artificial metalloenzymes have received increasing attention over the last decade as a possible solution to unaddressed challenges in synthetic organic chemistry. Whereas traditional transition-metal catalysts typically only take advantage of the first coordination sphere to control reactivity and selectivity, artificial metalloenzymes can modulate both the first and second coordination spheres. This difference can manifest itself in reactivity profiles that can be truly unique to artificial metalloenzymes. This Review summarizes attempts to modulate the second coordination sphere of artificial metalloenzymes by using genetic modifications of the protein sequence. In doing so, successful attempts and creative solutions to address the challenges encountered are highlighted.

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“…Having identified the most promising bidentate ligand L^L 5, the corresponding ATHase [Biot-Cp*Ir(L^L 5)Cl]⊂Sav was assembled (Scheme 1c, d). [21][22][23] Next, we combined this ATHase with glucose dehydrogenase (GDH) to regenerate the consumed NAD(P)H. This enables the use of glucose as reductant, yielding NAD(P)H and glucono-δ-lactone (Scheme 3). The performance of the concurrent enzymatic cascade was optimized by screening Sav variants focusing on closelying positions S112 and K121.…”

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confidence: 99%

“…The performance of the concurrent enzymatic cascade was optimized by screening Sav variants focusing on closelying positions S112 and K121. 22 The reactions contained GDH (0.1 mg/ml), NAD + (1 mM, 2 mol%), glucose (60 mM, 120 mol%), Biot-Cp*Ir(L^L 5)Cl (50 µM, 0.1 mol%), Sav free binding sites (100 µM) and MDQ A (50 mM). Compared to [Biot-Cp*Ir(L^L 5)Cl], the conversion by [Biot-Cp*Ir(L^L 5)Cl]⊂Sav doubled, highlighting the shielding effect of the host protein (Scheme 3, entries 1 and 2).…”

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confidence: 99%

An NAD(P)H-Dependent Artificial Transfer Hydrogenase for Multienzymatic Cascades

2016

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Enzymes typically depend on either NAD(P)H or FADH 2 as hydride source for reduction purposes. In contrast, organometallic catalysts most often rely on isopropanol or formate to generate the reactive hydride moiety. Here we show that incorporation of a Cp*Ir cofactor possessing a biotin moiety and 4,7-dihydroxy-1,10-phenanthroline into streptavidin (Sav) yields an NAD(P)H-dependent artificial transfer hydrogenase (ATHase). This ATHase (0.1 mol%) catalyzes imine reduction with 1 mM NADPH (2 mol%), which can be concurrently regenerated by a glucose dehydrogenase (GDH) using only 1.2 equivalents glucose. A four enzyme cascade consisting of the ATHase, the GDH, a monoamine oxidase and a catalase leads to the production of enantiopure amines.The introduction of synthetic catalysts into a biological context is at the focus of current efforts in both synthetic-and chemical biology.1,2 Goals include i) the supplementation of existing-or engineered metabolic pathways, ii) decaging inactive forms of enzymes to trigger enzymatic cascades, 2,3 iii) shifting the redox equilibrium in cancer cells to induce apoptosis 4 or iv) produce fuels with the help of biological redox equivalents.3-5 Achieving high productivity of synthetic organometallic catalysts inside a living system remains challenging and progress is likely to be incremental. 2,6 In contrast, the combination of isolated enzymes and transition metal catalysts in carefully designed in vitro systems of modest complexity experiences increasing success. 7-9Transition metal-mediated formal hydride transfer occupies a prominent role in many of these initiatives. Remarkably, synthetic catalysts and enzymes have gained common ground for the conversion of ketones to alcohols, imines to amines (and vice versa), the reduction of activated double bonds and the racemisation of secondary alcohols and amines.10 A few isolated studies have shown that transition metal complexes can accept NAD(P)H as a hydride source. 5,11,12 To the best of our knowledge however, their concurrent use coupled with enzymatic processes, where the hydride is utilized in a productive fashion, have not been disclosed yet (Scheme 1a). We hypothesize that this may be traced back to the mutual deactivation of both transition metal catalysts and natural enzymes. To overcome this challenge, spatial separation of both catalytic partners has proven most effective. [13][14][15] In this context, artificial metalloenzymes (AMEs) have received increasing attention as alternative to both homogeneous catalysts and enzymes. 16 To test the versatility of AMEs for the implementation of enzymatic cascades, we reported on the compartmentalization of a biotinylated Cp*Ir-based transfer hydrogenation catalyst into streptavidin (Sav) variants (Scheme 1d).15 To drive the hydride transfer reactions at reasonable rates however, concentrations of formate in the molar range were required. 7,15,17 Such concentrations of formate may lead to inactivation of natural enzymes and are thus incompatible with in vivo application...

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A Dual Anchoring Strategy for the Localization and Activation of Artificial Metalloenzymes Based on the Biotin–Streptavidin Technology

Cited by 92 publications

References 32 publications

Expanding the Chemical Diversity in Artificial Imine Reductases Based on the Biotin–Streptavidin Technology

Expanding the Chemical Diversity in Artificial Imine Reductases Based on the Biotin–Streptavidin Technology

Genetic Optimization of Metalloenzymes: Enhancing Enzymes for Non‐Natural Reactions

An NAD(P)H-Dependent Artificial Transfer Hydrogenase for Multienzymatic Cascades

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