Evolution of Shape Complementarity and Catalytic Efficiency from a Primordial Antibody Template

Xu, Jian; Deng, Qiaolin; Chen, Jiangang; Houk, K. N.; Bartek, J.; Hilvert, Donald; Wilson, Ian A.

doi:10.1126/science.286.5448.2345

Cited by 116 publications

(121 citation statements)

References 40 publications

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“…Figure 12 shows a typical antibody binding pocket, both with and without a bound hapten. [32] The dimensions of the Figure 9. Examples of hosts for organic molecules in non-aqueous solution.…”

Section: Antibody-antigen Complexesmentioning

confidence: 99%

Binding Affinities of Host–Guest, Protein–Ligand, and Protein–Transition‐State Complexes

et al. 2003

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The affinities of hosts—ranging from small synthetic cavitands to large proteins—for organic molecules are well documented. The average association constants for the binding of organic molecules by cyclodextrins, synthetic hosts, and albumins in water, as well as of catalytic antibodies or enzymes for substrates are 103.5±2.5 M−1. Binding affinities are elevated to 108±2 M−1 for the complexation of transition states and biological antigens by antibodies or inhibitors by enzymes, and to 1016±4 M−1 for transition states with enzymes. The origins of the distributions of association constants observed for the broad range of host–guest systems are explored in this Review, and typical approaches to compute and analyze host–guest binding in solution are discussed. In many classes of complexes a rough correlation is found between the binding affinity and the surface area that is buried upon complexation. Enzymes transcend this effect and achieve transition‐state binding much greater than is expected from the surface areas.

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Section: Antibody-antigen Complexesmentioning

confidence: 99%

Binding Affinities of Host–Guest, Protein–Ligand, and Protein–Transition‐State Complexes

et al. 2003

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“…In case of 10F11, the bound ligands are clearly visible in the electron density maps, allowing unequivocal identification of the catalytic pocket (Fig. 2), which is mainly hydrophobic as in other Diels-Alderase antibodies (11)(12)(13)(14)(15). The ligands are deeply buried in this pocket, which is formed by Tyr H53 , Tyr H58 , Ser H100 , Phe H101 , Trp H104 ͞Tyr H104 , Tyr L37 , Gly L96 , and Phe L99 (Fig.…”

Section: Resultsmentioning

confidence: 97%

“…3). Contrary to Diels-Alderase antibody 1E9 (11), where the fit between protein and ligand is so snug that no interfacial cavities are discernable, the binding pockets of 9D9 and 10F11 seem large enough (11 Å deep, 14 Å wide) to accommodate two to three water molecules beside the ligands. Ligand recognition is achieved through multiple van der Waals contacts.…”

Section: Resultsmentioning

confidence: 99%

“…Thus, immunization is carried out against a hapten with a molecular shape and charge distribution resembling those of the transition state as closely as possible. Although charges play a key role to mimic transition states of ionic reactions such as acidic or basic hydrolytic reactions, the shape of the transition state analogue has been particularly critical for the Diels-Alder reaction, for which several catalytic antibodies have been reported (5-10) and crystallized (11)(12)(13)(14)(15). Aromatic residues have been observed recurrently in the binding pockets of Diels-Alderase antibodies.…”

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confidence: 99%

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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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“…One substrate is physically tethered to the members of an RNA library, and active sequences are enriched by selection for the formation of a product, which remains tethered to the RNA and allows either selective amplification or affinity purification of the RNA. The crystal structures of a natural Diels-Alderase enzyme (11), a Diels-Alderase ribozyme (12), and several catalytic antibodies have now been elucidated (13)(14)(15), providing a structural framework for comparing the mechanisms of these different biopolymers.…”

mentioning

confidence: 99%

Selection of ribozymes that catalyse multiple-turnover Diels–Alder cycloadditions by using in vitro compartmentalization

Agresti

Kelly²,

Jäschke³

et al. 2005

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

110

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In vitro compartmentalization (IVC) has previously been used to evolve protein enzymes. Here, we demonstrate how IVC can be applied to select RNA enzymes (ribozymes) for a property that has previously been unselectable: true intermolecular catalysis. Libraries containing 10 11 ribozyme genes are compartmentalized in the aqueous droplets of a water-in-oil emulsion, such that most droplets contain no more than one gene, and transcribed in situ. By coencapsulating the gene, RNA, and the substrates͞products of the catalyzed reaction, ribozymes can be selected for all enzymatic properties: substrate recognition, product formation, rate acceleration, and turnover. Here we exploit the complementarity of IVC with systematic evolution of ligands by exponential enrichment (SELEX), which allows selection of larger libraries (>10 15 ) and for very small rate accelerations (kcat͞kuncat) but only selects for intramolecular single-turnover reactions. We selected Ϸ10 14 random RNAs for Diels-Alderase activity with five rounds of SELEX, then six to nine rounds with IVC. All selected ribozymes catalyzed the Diels-Alder reaction in a truly bimolecular fashion and with multiple turnover. Nearly all ribozymes selected by using eleven rounds of SELEX alone contain a common catalytic motif. Selecting with SELEX then IVC gave ribozymes with significant sequence variations in this catalytic motif and ribozymes with completely novel motifs. Interestingly, the catalytic properties of all of the selected ribozymes were quite similar. The ribozymes are strongly product inhibited, consistent with the Diels-Alder transition state closely resembling the product. More efficient Diels-Alderases may need to catalyze a second reaction that transforms the product and prevents product inhibition.emulsion ͉ RNA ͉ intermolecular catalysis T he Diels-Alder [4 ϩ 2] cycloaddition reaction (1) between a 1,3 diene and an alkene dienophile is one of the most useful and important in synthetic chemistry, because it allows the formation of six-membered rings by making two simultaneous C-C bonds and at the same time generates up to four chiral centers (2). However, nature rarely seems to use this mechanism for C-C bond formation and instead prefers other mechanisms such as the aldol condensation reaction. Although Ͼ100 natural products have been proposed to be Diels-Alder cycloadducts (3), direct evidence from biosynthetic studies with purified or partially purified enzymes is available for only three naturally occurring Diels-Alder reactions (4).However, several antibodies and RNAs that catalyze Diels-Alder reactions have been generated in the laboratory. The DielsAlderase antibodies were raised against a hapten that mimicked either the Diels-Alder adduct or the transition state of the desired reactants (reviewed in ref. 5). Diels-Alderase ribozymes with both pyridyl-modified (6, 7) and unmodified RNA (8) were generated by using a variation on systematic evolution of ligands by exponential enrichment (SELEX) (9, 10). One substrate is physically tethered to the...

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Evolution of Shape Complementarity and Catalytic Efficiency from a Primordial Antibody Template

Cited by 116 publications

References 40 publications

Binding Affinities of Host–Guest, Protein–Ligand, and Protein–Transition‐State Complexes

Binding Affinities of Host–Guest, Protein–Ligand, and Protein–Transition‐State Complexes

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

Selection of ribozymes that catalyse multiple-turnover Diels–Alder cycloadditions by using in vitro compartmentalization

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