title: Molecular analysis of antimicrobial agent translocation through the membrane porin BpsOmp38 from an ultraresistant Burkholderia pseudomallei strain.
abstract: Burkholderia pseudomallei (Bps) is a Gram-negative bacterium that causes melioidosis, an infectious disease of animals and humans common in northern and north-eastern parts of Thailand. Successful treatment of melioidosis is difficult due to intrinsic resistance of Bps to various antibacterial agents. It has been suggested that the antimicrobial resistance of this organism may result from poor permeability of the active compounds through porin channels located in the outer membrane (OM) of the bacterium. In previous work, a 38-kDa protein, named "BpsOmp38", was isolated from the OM of Bps. A topology prediction and liposome-swelling assay suggested that BpsOmp38 comprises a β-barrel structure and acts as a general diffusion porin. The present study employed black lipid membrane (BLM) reconstitution to demonstrate the single-channel conductance of the trimeric BpsOmp38 to be 2.7±0.3 nS in 1M KCl. High-time resolution BLM measurements displayed ion current blockages of seven antimicrobial agents in a concentration-dependent manner with the translocation on-rate (kon) following the order: norfloxacin≫ertapenem>ceftazidime>cefepime>imipenem>meropenem>penicillin G. The dwell time of a selected antimicrobial agent (ertapenem) decayed exponentially with increasing temperature. The energy barrier for the ertapenem binding to the affinity site inside the BpsOmp38 channel was estimated from the Arrhenius plot to be 12 kT and for the ertapenem release to be 13 kT at +100 mV. The BLM data obtained from this study provide the first insight into antimicrobial agent translocation through the BpsOmp38 channel.
authors.affiliation: Biochemistry-Electrochemistry Research Unit, School of Chemistry, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand. wipa@sut.ac.th
authors.forename: Wipa
authors.initials: W
authors.lastname: Suginta
authors.forename: Kozhinjampara R
authors.initials: KR
authors.lastname: Mahendran
authors.forename: Watcharin
authors.initials: W
authors.lastname: Chumjan
authors.forename: Eric
authors.initials: E
authors.lastname: Hajjar
authors.forename: Albert
authors.initials: A
authors.lastname: Schulte
authors.forename: Mathias
authors.initials: M
authors.lastname: Winterhalter
authors.forename: Helge
authors.initials: H
authors.lastname: Weingart
mesh_terms: D000595:Amino Acid Sequence; D000890:Anti-Infective Agents / Q000378:metabolism* / Q000493:pharmacokinetics; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport; D016957:Burkholderia pseudomallei / Q000235:genetics / Q000378:metabolism*; D000077723:Cefepime; D002442:Ceftazidime / Q000378:metabolism / Q000493:pharmacokinetics; D002511:Cephalosporins / Q000378:metabolism / Q000493:pharmacokinetics; D000077727:Ertapenem; D006801:Humans; D015378:Imipenem / Q000378:metabolism / Q000493:pharmacokinetics; D007700:Kinetics; D008051:Lipid Bilayers / Q000737:chemistry / Q000378:metabolism; D008554:Melioidosis / Q000382:microbiology; D000077731:Meropenem; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009643:Norfloxacin / Q000378:metabolism / Q000493:pharmacokinetics; D010400:Penicillin G / Q000378:metabolism / Q000493:pharmacokinetics; D010743:Phospholipids / Q000737:chemistry / Q000378:metabolism; D018272:Porins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D013845:Thienamycins / Q000378:metabolism / Q000493:pharmacokinetics; D047090:beta-Lactams / Q000378:metabolism / Q000493:pharmacokinetics
chemical_list: D000890:Anti-Infective Agents; D001425:Bacterial Outer Membrane Proteins; D002511:Cephalosporins; D008051:Lipid Bilayers; D010743:Phospholipids; D018272:Porins; D013845:Thienamycins; D047090:beta-Lactams; D015378:Imipenem; D000077723:Cefepime; D002442:Ceftazidime; D000077731:Meropenem; D000077727:Ertapenem; D009643:Norfloxacin; D010400:Penicillin G