5'-Methylthioadenosine/S -adenosylhomocysteine nucleosidase (MTAN) is a bacterial enzyme that is involved in quorum sensing pathways and a potential target for the design of antibiotics. Kinetic isotope effects (KIEs) and computer modeling using density functional theory were used to determine the transition state of N. meningitides MTAN. KIEs were measured for hydrolysis of 5'- methylthioadenosine (MTA) and were corrected for forward commitment to catalysis to obtain intrinsic KIEs for [1'-H-3], [9-N-15], [1'-C-14], [2'-H-3], [4'-H-3], [5'-H-3(2)], and [Me-H-3(3)] MTAs. KIEs support an early S(N)1 transition state unlike the dissociative transition states of other MTANs. Computational modeling of intrinsic KIEs places the leaving group adenine at 1.68 angstrom from the anomeric carbon, a Pauling bond order of 0.50, and no participation of the hydroxide nucleophile. The N7 of adenine is protonated and the leaving group is cationic owing to the significant bond order to the C1'-N9 bond at the transition state. The 3'-OH is not strongly polarized or ionized at the transition state, unlike other MTANs. The early transition state of N. meningitides MTAN is supported by its inhibition with transition-state analogues.