BN/CC isosteric compounds as enzyme inhibitors: N- and B-ethyl-1,2-azaborine inhibit ethylbenzene hydroxylation as nonconvertible substrate analogues

Abstract

Good substrate gone bad! BN/CC isosterism of ethylbenzene leads to N-ethyl-1,2-azaborine and B-ethyl-1,2-azaborine. In contrast to ethylbenzene, which is the substrate for ethylbenzene dehydrogenase (EbDH), N-ethyl-1,2-azaborine (see scheme; Fc=Ferricenium tetrafluoroborate) and B-ethyl-1,2-azaborine are strong inhibitors of EbDH. Thus, the changes provided by BN/CC isosterism can lead to new biochemical reactivity.

Figure 1

A: Nonlinear plots of the remaining ethylbenzene-dependent activity of EbDH when preincubated with different concentrations of N-ethyl-1,2-azaborine (circles) and B-ethyl-1,2-azaborine (squares). B: Double reciprocal plot of ethylbenzene-dependent enzymatic activity obtained when preincubated with different concentrations of N-ethyl-1,2-azaborine: 0 μm (triangles), 2.5 μm (diamonds), 10 μm (circles) 25 μm (squares). The kinetic data were fitted using the model for mixed inhibition^[17] (see also Supporting Information).

Figure 2. Overlay of N-ethyl-1,2-azaborine (red), B-ethyl-1,2-azaborine (yellow), and ethylbenzene (gray) modeled into the EbDH active site. The protein surface is given in grades of hydrophobicity (brown= hydrophobic, blue=hydrophilic). The view leads from the substrate channel towards the molybdenum cofactor. Note that force-field parameters had to be estimated for modeling the 1,2-azaborines, which may cause some deviations from the actual binding mode (see Supporting Information)

Scheme 1

Effect of BN/CC isosterism on enzyme reactivity.