Fig. 4: Investigation of the catalytic mechanism of BHMeHis1.8.
From: A non-canonical nucleophile unlocks a new mechanistic pathway in a designed enzyme
A KIE plots showing changes in reaction rate using either 2-cyclohexen-1-one (1, black lines) or 2-deuterocyclohex-2-en-1-one (S2, red lines) for BHMeHis1.8 and BHMeHis1.8 Glu26Gln. Reactions were performed using 1 or S2 (25 mM), 2 (2 mM) in PBS pH 7.0 with 3% (v/v) MeCN as cosolvent. Reactions were performed in triplicate. Source data are provided as a Source Data file. B Representative MD snapshot of BHMeHis1.8:Int2 complex with a protonated glutamic acid Glu(H)26 (model A) from a 500 ns simulation. Int2 (black) and key amino acid residues (blue) are shown in ball and stick representation with hydrogen bonds shown as black dashed lines. C Proposed mechanisms of BHMeHis1.8 showing a concerted (top) and a stepwise (bottom) proton transfer from Int2 to Int3 mediated by Glu(H)26.
