Two classification schemes for beta-lactamases are currently in use. The molecular classification is based on the amino acid sequence and divides beta-lactamases into class A, C, and D enzymes which utilize serine for beta-lactam hydrolysis and class B metalloenzymes which require divalent zinc ions for substrate hydrolysis. The functional classification scheme updated herein is based on the 1995 proposal by Bush et al. (K. Bush, G. A. Jacoby, and A. A. Medeiros, Antimicrob. Agents Chemother. 39:1211-1233, 1995). It takes into account substrate and inhibitor profiles in an attempt to group the enzymes in ways that can be correlated with their phenotype in clinical isolates. Major groupings generally correlate with the more broadly based molecular classification. The updated system includes group 1 (class C) cephalosporinases; group 2 (classes A and D) broad-spectrum, inhibitor-resistant, and extended-spectrum beta-lactamases and serine carbapenemases; and group 3 metallo-beta-lactamases. Several new subgroups of each of the major groups are described, based on specific attributes of individual enzymes. A list of attributes is also suggested for the description of a new beta-lactamase, including the requisite microbiological properties, substrate and inhibitor profiles, and molecular sequence data that provide an adequate characterization for a new beta-lactam-hydrolyzing enzyme.