O-Aryloxycarbonyl hydroxamates represent a new class of beta-lactamase inhibitors. N-Benzyloxycarbonyl-O-(phenoxycarbonyl) hydroxylamine, for example, inactivates the class C Enterobacter cloacae P99 beta-lactamase with a rate constant of 6.1 x 10(3) s(-1) M-1; approximately two turnover events accompany the inhibition. N-Benzyloxycarbonyl-O-[(3-carboxyphenoxy)carbonyl] hydroxylamine is comparably effective. These compounds also inactivate the class A TEM beta-lactamase. A crystal structure of the inactivated AmpC enzyme, another class C beta-lactamase, reveals that the active site has become cross-linked by a carbamate bridge spanning Ser64, the active site nucleophile, and Lys315, a conserved active site residue.