A Nitroxyl Synthase Catalytic Antibody

Bahr, Nicolaus; Güller, Rolf; Reymond, Jean‐Louis; Lerner, Richard A.

doi:10.1021/ja9538694

Cited by 48 publications

(39 citation statements)

References 37 publications

(21 reference statements)

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“…It is 30-to 300-fold more active than catalytic antibodies that promote the same [4+2] cycloaddition (9,11), and it outperforms the best artificial enzymes for other Diels-Alder reactions, including antibody catalysts (7)(8)(9)(10)(11)(12), ribozymes (13)(14)(15)(16), and artificial metalloenzymes (17,18) (Fig. 5D and Table S3).…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Impact of scaffold rigidity on the design and evolution of an artificial Diels-Alderase

Preiswerk

Beck

Schulz

et al. 2014

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

119

101

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By combining targeted mutagenesis, computational refinement, and directed evolution, a modestly active, computationally designed DielsAlderase was converted into the most proficient biocatalyst for [4+2] cycloadditions known. The high stereoselectivity and minimal product inhibition of the evolved enzyme enabled preparative scale synthesis of a single product diastereomer. X-ray crystallography of the enzyme-product complex shows that the molecular changes introduced over the course of optimization, including addition of a lid structure, gradually reshaped the pocket for more effective substrate preorganization and transition state stabilization. The good overall agreement between the experimental structure and the original design model with respect to the orientations of both the bound product and the catalytic side chains contrasts with other computationally designed enzymes. Because design accuracy appears to correlate with scaffold rigidity, improved control over backbone conformation will likely be the key to future efforts to design more efficient enzymes for diverse chemical reactions.biocatalysis | computational enzyme design | Diels-Alder reaction | laboratory evolution | enzyme mechanism

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Section: Discussionmentioning

confidence: 99%

“…A range of biocatalysts for both normal and inverse electron-demand Diels-Alder reactions has been elicited in response to immunogenic transition-state analogs (7)(8)(9)(10)(11)(12). Analogous nucleic acid-based catalysts have been isolated from large random libraries with powerful in vitro selection techniques (13)(14)(15)(16).…”

mentioning

confidence: 99%

Impact of scaffold rigidity on the design and evolution of an artificial Diels-Alderase

Preiswerk

Beck

Schulz

et al. 2014

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

119

101

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“…A similar catalytic role of hydrogen bonds has been argued for Diels-Alderase antibodies 1E9 (11) and 39A11 (12). In the slower reacting substrate 1b, the H-bonding interaction would occur with the nonbonding electron pair of the bridge's NH group, which should rather slow down the retro-Diels-Alder reaction if one considers that the retro-Diels-Alder reaction does not occur in the protonated forms of the substrates (16,17). Therefore, the participation of these hydrogen bonds could at most explain the 10-fold reactivity difference between the regioisomeric substrates 1a and 1b.…”

Section: Fig 2 Sequence Alignment and Comparison Of The Cdr H (A) Amentioning

confidence: 59%

“…Aromatic residues have been observed recurrently in the binding pockets of Diels-Alderase antibodies. We have used x-ray crystallography to identify a similar aromatic residue in the binding pocket of the retro-Diels-Alder catalytic antibodies 10F11 (16) and 9D9 (17). This aromatic residue is conserved as well in the third retro-Diels-Alder antibody 27C5.…”

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confidence: 99%

A structural basis for the activity of retro-Diels–Alder catalytic antibodies: Evidence for a catalytic aromatic residue

Hugot

Bensel

Vogel

et al. 2002

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

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The nitroxyl synthase catalytic antibodies 10F11, 9D9, and 27C5 catalyze the release of nitroxyl from a bicyclic pro-drug by accelerating a retro-Diels-Alder reaction. The Fabs (antigen-binding fragments) of these three catalytic antibodies were cloned and sequenced. Fab 9D9 was crystallized in the apo-form and in complex with one transition state analogue of the reaction. Crystal structures of Fab 10F11 in complex with ligands mimicking substrate, transition state, and product have been determined at resolutions ranging from 1.8 to 2.3 Å. Antibodies 9D9 and 10F11 show increased shape complementarity (as quantified by the program SC) to the hapten and to a modeled transition state as compared with substrate and product. The shape complementarity is mediated to a large extent by an aromatic residue (tyrosine or tryptophan) at the bottom of the hydrophobic active pocket, which undergoes -stacking interactions with the aromatic rings of the ligands. Another factor contributing to the different reactivity of the regioisomers probably arises because of hydrogen-bonding interactions between the nitroxyl bridge and the backbone amide of PheH101 and possibly a conserved water molecule.

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“…16,17 However, Pd(OAc) 2 /HCO 2 K in DMF at 50 °C selectively reduces 8 and 9 to 10 in quantitative yield. 18 Condensation of 10 with NH 2 OH hydrochloride/KOH gives hydroxamic acid (11) in 85% yield at −10 °C.…”

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confidence: 99%

Grignard Reagent-Mediated Conversion of an Acyl Nitroso-anthracene Cycloadduct to a Nitrone

2006

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An intramolecular hetero-Diels-Alder cycloadduct of an acyl nitroso compound and a 9, 10-dimethyl anthracene derivative was prepared as a potential nitroxyl (HNO) donor. This compound did not release HNO under any of the conditions tested. Treatment of this cycloadduct with excess MeMgCl resulted in the formation of a nitrone, whose structure was confirmed by X-ray crystallography. A mechanism where MeMgCl acts as a nucleophile, strong base and Lewis acid possibly explains the formation of this product.The cycloadducts of acyl nitroso compounds and 9,10-dimethylanthracene (1, Scheme 1) undergo thermal decomposition through retro-Diels-Alder reactions to produce acyl nitroso compounds (2, Scheme 1) under non-oxidative conditions and relatively mild temperatures (40-100°C). [1][2][3][4] Decomposition of these compounds provides a particularly clean method for the formation of acyl nitroso compounds, which are highly reactive N-O heterodienophiles. Acyl nitroso compounds also hydrolyze to release nitroxyl (HNO), the one-electron reduced form of the biologically important messenger molecule, nitric oxide (NO). [5][6][7][8][9] Nitroxyl has drawn considerable recent interest as it demonstrates distinct biological effects to NO in various systems. 10,11 Such activities and the inherent chemical instability of HNO also have focused efforts on the development of chemically and mechanistically unique nitroxyl releasing systems.Our recent studies show HNO release from functionalized N-hydroxyurea-derived acyl nitroso-9, 10-dimethylanthracene cycloadducts (1, R = -NHR) through the pathway depicted in Scheme 1. 12,13 Generation of an equivalent of the water-insoluble and immunotoxic 9, 10-dimethylanthracene (3, 9, 10-DMA) in this sequence remains a major limitation of this method of HNO formation. 14 The introduction of functional groups that act as hydrogenbond donors or acceptors on the 9, 10-DMA portion of these molecules should increase water solubility and reduce toxicity. 15 Compound 4, the product of an intramolecular heteroDiels-Alder reaction of acyl nitroso compound (5), fulfills this requirement by producing the carboxylic acid (6) after retro-Diels-Alder reaction and hydrolysis (Scheme 1). We wish to report the preparation of 4, its ability to act as an HNO donor and a unique molecular rearrangement upon treatment with an excess amount of Grignard reagent.Scheme 2 depicts the synthesis of the target compound 4. Horner-Wadsworth-Emmons olefination of commercially available 10-methylanthracene-9-carboxaldehyde (7) yields a mixture of the α, β-unsaturated esters 8 and 9 (8:9 = 5.8:1, 92% overall yield, Scheme 2). Reduction of 8 and 9 proved difficult with NaBH 4 /NiCl 2 and hydrogenation on Pd/C yielding mixtures as a result of the concurrent reduction of the anthracene moiety. 16,17 However, Pd(OAc) 2 /HCO 2 K in DMF at 50 °C selectively reduces 8 and 9 to 10 in quantitative yield. 18 Condensation of 10 with NH 2 OH hydrochloride/KOH gives hydroxamic acid (11) in 85% yield at −10 °C. 19 Tetra-n-butylammoniu...

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A Nitroxyl Synthase Catalytic Antibody

Cited by 48 publications

References 37 publications

Impact of scaffold rigidity on the design and evolution of an artificial Diels-Alderase

Impact of scaffold rigidity on the design and evolution of an artificial Diels-Alderase

A structural basis for the activity of retro-Diels–Alder catalytic antibodies: Evidence for a catalytic aromatic residue

Grignard Reagent-Mediated Conversion of an Acyl Nitroso-anthracene Cycloadduct to a Nitrone

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