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Publication Abstract from PubMed

Combinations of beta-lactams with clavulanate are currently investigated for tuberculosis treatment. Since Mycobacterium tuberculosis produces a broad spectrum beta-lactamase, BlaC, the success of this approach could be compromised by the emergence of clavulanate-resistant variants, as observed for inhibitor resistant TEM variants in enterobacteria. Previous analyses based on site-directed mutagenesis of BlaC have led to the conclusion that this risk was limited. Here, we used a different approach based on determination of the crystal structure of beta-lactamase BlaMab of Mycobacterium abscessus, which efficiently hydrolyses clavulanate. Comparison of BlaMab and BlaC allowed for structure-assisted site-directed mutagenesis of BlaC and identification of the G132N substitution that was sufficient to switch interaction of BlaC with clavulanate from irreversible inactivation to efficient hydrolysis. The substitution, which restored the canonical SDN motif (SDG-->SDN), allowed for efficient hydrolysis of clavulanate, with a more than 104 increase in kcat (0.41 s-1), without affecting hydrolysis of other beta-lactams. Mass spectrometry revealed that acylation of BlaC and of its G132N variant by clavulanate follows similar paths involving sequential formation of two acylenzymes. Decarboxylation of the first acylenzyme results in a stable secondary acylenzyme in BlaC, whereas hydrolysis occurs in the G132N variant. The SDN/SDG polymorphism defines two mycobacterial lineages comprising rapidly and slowly growing species, respectively. Together, these results suggest that the efficacy of beta-lactam-clavulanate combinations may be limited by emergence of resistance. beta-lactams active without clavulanate, such as faropenem, should be prioritized for development of new therapies.

Hydrolysis of clavulanate by Mycobacterium tuberculosis beta-lactamase BlaC harboring a canonic SDN motif.,Soroka D, Li de la Sierra-Gallay I, Dubee V, Triboulet S, van Tilbeurgh H, Compain F, Ballell L, Barros D, Mainardi JL, Hugonnet JE, Arthur M Antimicrob Agents Chemother. 2015 Jul 6. pii: AAC.00598-15. PMID:26149997^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.