title: The Carboxyl Terminus of Eremomycin Facilitates Binding to the Non-d-Ala-d-Ala Segment of the Peptidoglycan Pentapeptide Stem.
abstract: Glycopeptide antibiotics inhibit cell wall biosynthesis in Gram-positive bacteria by targeting the peptidoglycan (PG) pentapeptide stem structure (l-Ala-d-iso-Gln-l-Lys-d-Ala-d-Ala). Structures of the glycopeptide complexed with a PG stem mimic have shown that the d-Ala-d-Ala segment is the primary drug binding site; however, biochemical evidence suggests that the glycopeptide-PG interaction involves more than d-Ala-d-Ala binding. Interactions of the glycopeptide with the non-d-Ala-d-Ala segment of the PG stem were investigated using solid-state nuclear magnetic resonance (NMR). LCTA-1421, a double (15)N-enriched eremomycin derivative with a C-terminal [(15)N]amide and [(15)N]Asn amide, was complexed with whole cells of Staphylococcus aureus grown in a defined medium containing l-[3-(13)C]Ala and d-[1-(13)C]Ala in the presence of alanine racemase inhibitor alaphosphin. (13)C{(15)N} and (15)N{(13)C} rotational-echo double-resonance (REDOR) NMR measurements determined the (13)C-(15)N internuclear distances between the [(15)N]Asn amide of LCTA-1421 and the (13)C atoms of the bound d-[1-(13)C]Ala-d-[1-(13)C]Ala to be 5.1 and 4.8 Å, respectively. These measurements also determined the distance from the C-terminal [(15)N]amide of LCTA-1421 to the l-[3-(13)C]Ala of PG to be 3.5 Å. The measured REDOR distance constraints position the C-terminus of the glycopeptide in the proximity of the l-Ala of the PG, suggesting that the C-terminus of the glycopeptide interacts near the l-Ala segment of the PG stem. In vivo REDOR measurements provided structural insight into how C-terminally modified glycopeptide antibiotics operate.
authors.affiliation: Department of Chemistry and Biochemistry, Baylor University , Waco, Texas 76706, United States.
authors.forename: James
authors.initials: J
authors.lastname: Chang
authors.affiliation: Department of Chemistry, Center for Drug Discovery, Design, and Delivery (CD4), Center for Scientific Computation, Southern Methodist University , Dallas, Texas 75275, United States.
authors.forename: Hongyu
authors.initials: H
authors.lastname: Zhou
authors.affiliation: Gause Institute of New Antibiotics , Moscow, Russian Federation.
authors.forename: Maria
authors.initials: M
authors.lastname: Preobrazhenskaya
authors.affiliation: Department of Chemistry, Center for Drug Discovery, Design, and Delivery (CD4), Center for Scientific Computation, Southern Methodist University , Dallas, Texas 75275, United States.
authors.forename: Peng
authors.initials: P
authors.lastname: Tao
authors.affiliation: Department of Chemistry and Biochemistry, Baylor University , Waco, Texas 76706, United States.
authors.forename: Sung Joon
authors.initials: SJ
authors.lastname: Kim
mesh_terms: D000409:Alanine / Q000737:chemistry / Q000378:metabolism*; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001665:Binding Sites; D002473:Cell Wall / Q000187:drug effects / Q000378:metabolism*; D006020:Glycopeptides / Q000494:pharmacology*; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008968:Molecular Conformation; D056004:Molecular Dynamics Simulation; D010457:Peptidoglycan / Q000737:chemistry / Q000378:metabolism*; D013211:Staphylococcus aureus / Q000187:drug effects / Q000254:growth & development / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D006020:Glycopeptides; D010457:Peptidoglycan; C053759:eremomycin; D000409:Alanine