The genomes of both slow-growing and rapidly-growing mycobacterial species encode multiple proteins with the L,D-transpeptidase (Ldt) sequence motif. While the reason for this redundancy is not well understood, studies indicate paralogous Ldts likely serve niche roles in maintaining and/or remodeling mycobacterial peptidoglycan (PG). Here, we report a phylogenetic study that examined 323 mycobacterial Ldts and demonstrate these enzymes cluster into six clades. Our analysis revealed a variably represented class of Ldts that segregate into a monophyletic clade containing Mycobacterium smegmatis (Msm) LdtF. LdtF is closely related to class 2 Ldts like LdtB, and the crystal structures of other class 2 Ldts have been reported. We built a homology model of Msm LdtF in order to understand structural differences among this unique enzyme class. Our model indicates class 6 Ldts have structurally divergent catalytic domains containing a 10-residue insertion near the active site. Biochemical studies on Msm Ldts show that Ldts belonging to classes 1–4 are preferentially acylated by the penem faropenem in a competition assay, whereas the carbapenem meropenem preferentially acylates LdtF, a class 6 Ldt. Time course experiments reveal that the faropenem adduct is hydrolyzed from LdtF, whereas the LdtB-faropenem adduct is stable. Thus, our data demonstrate an evolutionary basis for mycobacterial Ldt multiplicity that supports the notion that paralogous Ldts serve physiological roles that are not redundant, and suggests class 6 Ldts can be selectively targeted by specific carbapenem antibiotics.

Phylogenetic and biochemical analyses of mycobacterial L,D-transpeptidases suggest paralogous enzymes can be selectively targeted by carbapenem antibiotics

Macromolecular Chemistry: The Second Century

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ACS Spring 2020 National Meeting & Expo

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.