β-Lactamase enzymes have attracted substential medical attention from researchers and clinicians because of their clinical, ecological, and evolutionary interest. Here, we present a comprehensive online database of β-lactamase enzymes. The current database is manually curated and incorporates the primary amino acid sequences, closest structural information in an external structure database (the Protein Data Bank [PDB]) and the functional profiles and phylogenetic trees of the four molecular classes (A, B, C, and D) of β-lactamases. The functional profiles are presented according to the MICs and kinetic parameters that make them more useful for the investigators. Here, a total of 1,147 β-lactam resistance genes are analyzed and described in the database. The database is implemented in MySQL and the related website is developed with Zend Framework 2 on an Apache server, supporting all major web browsers. Users can easily retrieve and visualize biologically important information using a set of efficient queries from a graphical interface. This database is freely accessible at http://ifr48.timone.univ-mrs.fr/beta-lactamase/public/.