The msesMD simulation also appears to reproduce more subtle features of the unbiased simulations, in particular sampling of GlcNAc ring puckering. This can be seen from the free energy profiles as a function of Cremer-Pople pucker angle θ (Figure ). The msesMD simulations, as well as unbiased triplicate 1 s and 10 s MD trajectories sample the full spectrum of puckers, from 4 C 1 through skew-boat intermediates to 1 C 4 conformations, tracing similar profiles (Figure ). For the remaining sugar rings, the pucker free energy profiles obtained from msesMD broadly reflect those from unbiased MD albeit with some differences regarding the highest energy regions (Figure ); the greatest discrepancy is found for the high energy C 1 state of Neu5Ac where biased and unbiased estimates differ by ~2.5 kcal mol -1 (Figure ).