The marketed drug topiramate ( 1) is a moderate inhibitor of carbonic anhydrase-II (CA-II) ( K i or K d = 0.3-0.6 microM), whereas sulfamide cognate 2 is a comparatively weak inhibitor ( K i or K d = 25-650 microM). From an X-ray cocrystal structure of 2.CA-II, Winum et al. ( J. Med. Chem. 2006, 49, 7024) proposed that an adverse steric interaction between the C8 methyl group in 2 and Ala-65 of CA-II is responsible for the diminished CA-II inhibitory potency of 2. We performed a straightforward test of this Ala-65 effect by synthesizing and examining ligand 3, which lacks the offending (pro- S or C8) methyl substituent in 2. We also prepared and evaluated related sulfamides 5, 7, and 9. In a CA-II inhibition assay (4-nitrophenyl acetate), the K i for 3 was approximately 300 microM, indicating very weak inhibition, close to that for 2 (4NPA, K i = 340 microM). In a CA-II binding assay (ThermoFluor), the K d for 3 was >57 microM, indicating very weak binding, lower than the affinity of 2 ( K d = 25 microM). Our results draw into question the proposed steric interaction between the C8 methyl of 2 and Ala-65 of CA-II.