Routes to covalent catalysis by reactive selection for nascent protein nucleophiles

Andrey V. Reshetnyak, Maria Francesca Armentano, Natalia A. Ponomarenko, Domenica Vizzuso, Oxana M. Durova, Rustam Ziganshin, Marina Serebryakova, Vadim Govorun, Gennady Gololobov, Herbert C. Morse, Alain Friboulet, Sudesh P Makker, Alexander G. Gabibov, Alfonso Tramontano

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

Reactivity-based selection strategies have been used to enrich combinatorial libraries for encoded biocatalysts having revised substrate specificity or altered catalytic activity. This approach can also assist in artificial evolution of enzyme catalysis from protein templates without bias for predefined catalytic sites. The prevalence of covalent intermediates in enzymatic mechanisms suggests the universal utility of the covalent complex as the basis for selection. Covalent selection by phosphonate ester exchange was applied to a phage display library of antibody variable fragments (scFv) to sample the scope and mechanism of chemical reactivity in a naive molecular library. Selected scFv segregated into structurally related covalent and noncovalent binders. Clones that reacted covalently utilized tyrosine residues exclusively as the nucleophile. Two motifs were identified by structural analysis, recruiting distinct Tyr residues of the light chain. Most clones employed Tyr32 in CDR-L1, whereas a unique clone (A.17) reacted at Tyr36 in FR-L2. Enhanced phosphonylation kinetics and modest amidase activity of A.17 suggested a primitive catalytic site. Covalent selection may thus provide access to protein molecules that approximate an early apparatus for covalent catalysis.

Original languageEnglish (US)
Pages (from-to)16175-16182
Number of pages8
JournalJournal of the American Chemical Society
Volume129
Issue number51
DOIs
StatePublished - Dec 26 2007

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Nucleophiles
Catalysis
amidase
Clone Cells
Proteins
Organophosphonates
Chemical reactivity
Bacteriophages
Biocatalysts
Catalytic Domain
Enzymes
Antibodies
Structural analysis
Binders
Tyrosine
Catalyst activity
Esters
Immunoglobulin Fragments
Display devices
Substrate Specificity

ASJC Scopus subject areas

  • Chemistry(all)

Cite this

Reshetnyak, A. V., Armentano, M. F., Ponomarenko, N. A., Vizzuso, D., Durova, O. M., Ziganshin, R., ... Tramontano, A. (2007). Routes to covalent catalysis by reactive selection for nascent protein nucleophiles. Journal of the American Chemical Society, 129(51), 16175-16182. https://doi.org/10.1021/ja076528m

Routes to covalent catalysis by reactive selection for nascent protein nucleophiles. / Reshetnyak, Andrey V.; Armentano, Maria Francesca; Ponomarenko, Natalia A.; Vizzuso, Domenica; Durova, Oxana M.; Ziganshin, Rustam; Serebryakova, Marina; Govorun, Vadim; Gololobov, Gennady; Morse, Herbert C.; Friboulet, Alain; Makker, Sudesh P; Gabibov, Alexander G.; Tramontano, Alfonso.

In: Journal of the American Chemical Society, Vol. 129, No. 51, 26.12.2007, p. 16175-16182.

Research output: Contribution to journalArticle

Reshetnyak, AV, Armentano, MF, Ponomarenko, NA, Vizzuso, D, Durova, OM, Ziganshin, R, Serebryakova, M, Govorun, V, Gololobov, G, Morse, HC, Friboulet, A, Makker, SP, Gabibov, AG & Tramontano, A 2007, 'Routes to covalent catalysis by reactive selection for nascent protein nucleophiles', Journal of the American Chemical Society, vol. 129, no. 51, pp. 16175-16182. https://doi.org/10.1021/ja076528m
Reshetnyak AV, Armentano MF, Ponomarenko NA, Vizzuso D, Durova OM, Ziganshin R et al. Routes to covalent catalysis by reactive selection for nascent protein nucleophiles. Journal of the American Chemical Society. 2007 Dec 26;129(51):16175-16182. https://doi.org/10.1021/ja076528m
Reshetnyak, Andrey V. ; Armentano, Maria Francesca ; Ponomarenko, Natalia A. ; Vizzuso, Domenica ; Durova, Oxana M. ; Ziganshin, Rustam ; Serebryakova, Marina ; Govorun, Vadim ; Gololobov, Gennady ; Morse, Herbert C. ; Friboulet, Alain ; Makker, Sudesh P ; Gabibov, Alexander G. ; Tramontano, Alfonso. / Routes to covalent catalysis by reactive selection for nascent protein nucleophiles. In: Journal of the American Chemical Society. 2007 ; Vol. 129, No. 51. pp. 16175-16182.
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