Crystal structures of the cadmium‐ and mercury‐substituted metallo‐β‐lactamase from Bacteroides fragilis

• Published in: Protein science 1997-12, Vol.6 (12), p.2671-2676
• Main Authors: Concha, Nestor O., Herzberg, Osnat, Rasmussen, Beth A., Bush, Karen
• Format: Article
• Language: English
• Subjects: Bacteroides fragilis - enzymology; beta-Lactamases - chemistry; beta-Lactamases - metabolism; Binding Sites; binuclear metal center; Cadmium - chemistry; Cadmium - metabolism; cadmium‐binding; Catalysis; Crystallization; Crystallography, X-Ray; Mercury - chemistry; Mercury - metabolism; mercury‐binding; metallo‐@‐lactamase; Molecular Structure; Penicillin G - metabolism; Structure-Activity Relationship; X‐ray structure; Zinc - chemistry; Zinc - metabolism
• ISSN: 0961-8368; 1469-896X
• DOI: 10.1002/pro.5560061225

The metallo‐β‐lactamases require zinc or cadmium for hydrolyzing β‐lactam antibiotics and are inhibited by mercurial compounds. To date, there are no clinically useful inhibitors of this class of enzymes. The crystal structure of the Zn2+‐bound enzyme from Bacteroides fragilis contains a binuclear zinc center in the active site. A hydroxide, coordinated to both zinc atoms, is proposed as the moiety that mounts the nucleophilic attack on the carbonyl carbon atom of the β‐lactam ring. To study the metal coordination further, the crystal structures of a Cd2+‐bound enzyme and of an Hg2+‐soaked zinc‐containing enzyme have been determined at 2.I Å and 2.7 Å, respectively. Given the diffraction resolution, the Cd2+‐bound enzyme exhibits the same active‐site architecture as that of the Zn2+‐bound enzyme, consistent with the fact that both forms are enzymatically active. The 10‐fold reduction in activity of the Cd2+‐bound molecule compared with the Zn2+‐bound enzyme is attributed to fine differences in the charge distribution due to the difference in the ionic radii of the two metals. In contrast, in the Hg2+‐bound structure, one of the zinc ions, Zn2, was ejected, and the other zinc ion, Zn1, remained in the same site as in the 2‐Zn2+‐bound structure. Instead of the ejected zinc, a mercury ion binds between Cys 104 and Cys 181, 4.8 Å away from Zn1 and 3.9 Å away from the site where Zn2 is located in the 2‐Zn2+‐bound molecule. The perturbed binuclear metal cluster explains the inactivation of the enzyme by mercury compounds.