title: Loss of a Functionally and Structurally Distinct ld-Transpeptidase, LdtMt5, Compromises Cell Wall Integrity in Mycobacterium tuberculosis.
abstract: The final step of peptidoglycan (PG) biosynthesis in bacteria involves cross-linking of peptide side chains. This step in Mycobacterium tuberculosis is catalyzed by ld- and dd-transpeptidases that generate 3→3 and 4→3 transpeptide linkages, respectively. M. tuberculosis PG is predominantly 3→3 cross-linked, and LdtMt2 is the dominant ld-transpeptidase. There are four additional sequence paralogs of LdtMt2 encoded by the genome of this pathogen, and the reason for this apparent redundancy is unknown. Here, we studied one of the paralogs, LdtMt5, and found it to be structurally and functionally distinct. The structures of apo-LdtMt5 and its meropenem adduct presented here demonstrate that, despite overall architectural similarity to LdtMt2, the LdtMt5 active site has marked differences. The presence of a structurally divergent catalytic site and a proline-rich C-terminal subdomain suggest that this protein may have a distinct role in PG metabolism, perhaps involving other cell wall-anchored proteins. Furthermore, M. tuberculosis lacking a functional copy of LdtMt5 displayed aberrant growth and was more susceptible to killing by crystal violet, osmotic shock, and select carbapenem antibiotics. Therefore, we conclude that LdtMt5 is not a functionally redundant ld-transpeptidase, but rather it serves a unique and important role in maintaining the integrity of the M. tuberculosis cell wall.
authors: From the Taskforce to study Resistance Emergence and Antimicrobial development Technology (TREAT) and Division of Infectious Diseases, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21231.
authors: Leighanne A
authors: http://orcid.org/0000-0002-4121-5505
authors: LA
authors: Brammer Basta
authors: Structural Enzymology and Thermodynamics Group, Department of Biophysics and Biophysical Chemistry and.
authors: Anita
authors: A
authors: Ghosh
authors: Structural Enzymology and Thermodynamics Group, Department of Biophysics and Biophysical Chemistry and.
authors: Ying
authors: Y
authors: Pan
authors: National Synchrotron Light Source II, Brookhaven National Laboratory, Upton, New York 11973, and.
authors: Jean
authors: J
authors: Jakoncic
authors: Department of Chemistry, The Johns Hopkins University, Baltimore, Maryland 21218.
authors: Evan P
authors: EP
authors: Lloyd
authors: Department of Chemistry, The Johns Hopkins University, Baltimore, Maryland 21218.
authors: Craig A
authors: CA
authors: Townsend
authors: From the Taskforce to study Resistance Emergence and Antimicrobial development Technology (TREAT) and Division of Infectious Diseases, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21231, lamichhane@jhu.edu.
authors: Gyanu
authors: http://orcid.org/0000-0002-2214-0114
authors: G
authors: Lamichhane
authors: Structural Enzymology and Thermodynamics Group, Department of Biophysics and Biophysical Chemistry and Department of Neurology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, bianchet@jhmi.edu.
authors: Mario A
authors: http://orcid.org/0000-0001-9032-7549
authors: MA
authors: Bianchet
mesh_terms: D000595:Amino Acid Sequence; D002384:Catalysis; D020134:Catalytic Domain; D002473:Cell Wall / Q000502:physiology*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D057927:Hydrophobic and Hydrophilic Interactions; D008969:Molecular Sequence Data; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000378:metabolism / Q000502:physiology; D010457:Peptidoglycan / Q000378:metabolism; D010458:Peptidyl Transferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid
chemical_list: D010457:Peptidoglycan; D010458:Peptidyl Transferases
keywords: Mycobacterium tuberculosis; antibiotics; biosynthesis; cell wall; enzyme kinetics; enzyme structure; peptidoglycan