3.2 Mechanistic analysis of the SucP-catalyzed transglycosylation
Guided by the mathematical deductions from Figure 2, we carried out extensive initial-rate studies for Lm SucP and Ba SucP, to obtain experimental evidence suitable to achieve mechanistic discrimination. In particular, dependencies on [GlcX] were measured with glycerol present for \(v_{X}\) or absent for \(v_{X}\) and\(v_{H}\). Dependencies on [GOH] were measured for \(v_{X}\),\(v_{H}\) and \(\frac{v_{X}}{v_{H}}\). Donor substrate was saturating (sucrose: 20 mM, both enzymes; G1P: 50 mM, Lm SucP; 450 mM,Ba SucP). Using G1P, approximately half-saturating concentration was additionally applied (5 mM, Lm SucP; 50 mM; Ba SucP). The experimental data are shown together with the results of best-model fits in Figure 3 (Lm SucP) and Figure S2 (Ba SucP; Supporting Information).