Computational mutagenesis reveals the role of active-site tyrosine in stabilising a boat conformation for the substrate: QM/MM molecular dynamics studies of wild-type and mutant xylanases

Mahmoud E S Soliman, G D Ruggiero, J J R Pernia, I R Greig, Ian H Williams

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Abstract

Molecular dynamics simulations have been performed for non-covalent complexes of phenyl beta-xylobioside with the retaining endo-beta-1,4-xylanase from B. circulans (BCX) and its Tyr69Phe mutant using a hybrid QM/MM methodology. A trajectory initiated for the wild-type enzyme-substrate complex with the proximal xylose ring bound at the -1 subsite ( adjacent to the scissile glycosidic bond) in the C-4(1) chair conformation shows spontaneous transformation to the B-2,B-5 boat conformation, and potential of mean force calculations indicate that the boat is similar to 30 kJ mol(-1) lower in free energy than the chair. Analogous simulations for the mutant lacking one oxygen atom confirm the key role of Tyr69 in stabilizing the boat in preference to the 4C1 chair conformation, with a relative free energy difference of about 20 kJ mol(-1), by donating a hydrogen bond to the endocyclic oxygen of the proximal xylose ring. QM/MM MD simulations for phenyl beta-xyloside in water, with and without a propionate/propionic acid pair to mimic the catalytic glutamate/glutamic acid pair of the enzyme, show the 4C1 chair to be stable, although a hydrogen bond between the OH group at C2 of xylose and the propionate moiety seems to provide some stabilization for the B-2,B-5 conformation.
Original languageEnglish
Pages (from-to)460-468
Number of pages9
JournalOrganic and Biomolecular Chemistry
Volume7
Issue number3
Early online date5 Dec 2008
DOIs
Publication statusPublished - 7 Feb 2009

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