Extension of the hydrolysis spectrum of AmpC b-lactamase of Escherichia coli due to amino acid insertion in the H-10 helix—authors’ response

responsible for the extended substrate spectrum is the R2 loop (residues 289–310 according to the amino acid numbering system of P99; Figure 1). Another issue that we wish to highlight from our reading of the work of Mammeri et al. is a commonly made oversimplification that a decrease in Km indicates an increased affinity of a substrate for an enzyme. Because an increased k2/k3 ratio (not only decreased Km value) also yields an increased affinity, 6 the Km value is a complex constant that does not simply reflect the binding affinity, especially when the deacylation step (related to k2 and k3) is rate-limiting.