Recently we have reported that the kinetics of AcmB point to cholest-4-en-3-one (CHON) as a native substrate . AcmB differs from KstD1 by the presence of a 40 amino acid fragment (153-204) that was formerly referred to as a 'loop' but that in fact, forms a putative membraneassociated domain at the entrance of the active site (Figures and). Besides anchoring AcmB to the membrane and positioning the active site entrance toward it, this domain seems to be involved in the binding of substrates with extended C17 substituents, which stick out of the KstD1 active site. We assume that the presence of this fragment may be responsible for the unique substrate specificity of AcmB with respect to enzymes devoid of this sequence. To check this hypothesis, we decided to conduct a series of MD simulations and estimate the free energy of binding for AcmB:steroid complexes, as well as calculate the interaction energies of substrates with the whole protein and membrane-associated domain (Table ).