Microbial Efflux Pumps: Current Research
"does an excellent job" (JAVMA)
Caister Academic Press
Edward W. Yu, Qijing Zhang and Melissa H. Brown
Department of Chemistry and Department of Physics and Astronomy, Iowa State University, Ames, USA; Department of Veterinary Microbiology and Preventive Medicine, Iowa State University, Ames, USA; School of Biological Sciences, Flinders University, Adelaide, Australia; respectively
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Infectious diseases caused by bacteria remain a leading cause of death worldwide. Many of the antibiotics developed to combat bacterial infections have been rendered almost impotent due to the rapid evolution and spread of antibiotic resistance. A common and major resistance mechanism, the efflux system, enables bacteria to extrude structurally diverse antimicrobials, facilitating survival in toxic environments. The pumps also have important physiological functions, play major roles in bacterial pathogenesis and are distributed widely across diverse bacterial species. In addition a single species may harbour several different types of efflux systems: of these, active efflux has proven to be one of the most successful detoxification mechanisms used by both Gram-positive and -negative pathogens. Unravelling the intricacies of the microbial efflux systems is essential for the development of new strategies to overcome antimicrobial resistance. This has inspired a plethora of multidisciplinary research projects that have focused on the biochemistry, bioinformatics, structural and molecular biology of this fascinating field. With contributions from leading researchers in the field, this book reviews the most important current research and summarizes the most spectacular discoveries. Essential reading for all scientists with an interest in drug and antibiotic resistance in a range of different microorganisms.
"does an excellent job ... provides valuable information ... useful for research scientists ... pharmaceutical experts ... and for clinicians" from JAVMA (2013) 243: No 6.
Table of contents
1. Transmembrane Molecular Transporters Facilitating Export of Molecules from Cells and Organelles
Maksim A. Shlykov, Wei Hao Zheng, Eric Wang, Justin D. Nguyen, Milton H. Saier and Jr.
Transport systems catalyze the uptake and efflux of solutes and macromolecules from all living cells and organelles. In this chapter, we review multiple types of efflux systems responsible for the export of inorganic and organic cations and anions, metabolites, drugs, complex carbohydrates, lipids, proteins and nucleic acids. These types fall into three primary categories: simple channels, secondary carriers and primary active transporters. We select a number of well-characterized systems in order to illustrate the structures and functions of different types of transporters as well as their mechanisms of action. However, metabolite exporters, in contrast to uptake systems and drug and macromolecular exporters, are still poorly characterized. We anticipate that studies conducted in the near future will reveal a plethora of metabolite exporters while revealing relatively few novel systems for metabolite uptake.
2. Structures of Multidrug Efflux Pumps from the MFS, SMR, MATE, and ABC Transporter Families
Geoffrey Chang, Paul Szewczyk and Xiao He
Drug efflux pumps play an important role in multidrug resistance (MDR) affecting the treatment of several diseases like infection and cancer. In this chapter, we review some of the molecular structures of MDR transporters that have their substrate binding sites located in the lipid bilayer. These transporters include those from the major facilitator superfamily (MFS), small multidrug resistance (SMR) family, the multidrug and toxic-compound extrusion (MATE) family, and the ATP-binding cassette (ABC) family. From these models, common structural themes have emerged that are consistent with their function, including a hydrophobic substrate binding pocket comprising of several aromatic residues facilitating poly-specificity, V-shapes to allow access of substrate from the lipid bilayer, and distinct inward- and outward-facing conformations facilitating transport across the cell membrane.
3. The Machinery and Mechanism of Multidrug Efflux in Gram-negative Bacteria
Dijun Du, Henrietta Venter, Klaas M. Pos and Ben F. Luisi
The Gram-negative bacteria are enveloped with a protective double-layer of lipid membranes, across which they expel noxious compounds by active transmembrane transport. Several classes of membrane-spanning proteins mediate the transport and thereby confer the bacteria with the capacity to occupy hazardous ecological niches or to evade the cytotoxic effects of antimicrobial compounds. The proteins can be grouped into diverse structural families, but all drive transport uphill energetically, against concentration gradients, using the free energy of electrochemical gradient or of ATP hydrolysis. One group of trans-membrane proteins forms an assembly that spans the inner and outer membranes and the periplasm. The inner membrane component of the assembly recognizes substrates for transport and harnesses energy for the processes, whereas a major role of the periplasmic adapter is to connect the inner membrane component to the outer membrane channel to safeguard the transport of the captured substrate across the outer membrane. The outer membrane component has an adjustable aperture that may be opened through interaction with its partner pump proteins. Using the AcrA/AcrB/TolC assembly of Escherichia coli as a paradigm efflux machine, we describe the current knowledge of the pump components and speculate on how they might interact in an allosteric manner during the transport process.
4. Structure, Mechanism and Assembly of the Tripartite CusCBA Heavy-metal Efflux Complex
Sylvia V. Do, Chih-Chia Su, Feng Long, Hsiang-Ting Lei, Jani Reddy Bolla and Edward W. Yu
Gram-negative bacteria expel various toxic chemicals via tripartite efflux pumps belonging to the resistance-nodulation-cell division (RND) superfamily. These pumps span both the inner and outer membranes of the cell. The three components of these tripartite systems are an inner membrane, substrate-binding transporter (or pump); a periplasmic membrane fusion protein (or adaptor); and an outer membrane-anchored channel. These three efflux proteins interact in the periplasmic space to form the three-part complex. One such efflux system, CusCBA, is responsible for extruding Cu(I) and Ag(I) ions, which are biocides. We previously presented the crystal structures of both the inner membrane transporter CusA and membrane fusion protein CusB of the CusCBA tripartite efflux system from Escherichia coli. We also described the co-crystal structure of the CusBA adaptor-transporter, revealing that the trimeric CusA efflux pump assembles with six CusB protein molecules to form the complex CusB6-CusA3. Recently, we reported three different conformers of the crystal structures of CusBA-Cu(I), suggesting a mechanism on how Cu(I) binding initiates a sequence of conformational transitions in the transport cycle. Herein, we summarize the structural information of these efflux proteins, and present the accumulated evidence that this efflux system utilizes conserved methionine as well as charged amino acids to bind and export Cu(I) and Ag(I) ions.
5. RND-Efflux Pumps for Metal Cations
Dietrich H. Nies
This review will concentrate on metal-transporting RND-proteins in bacteria since such proteins have not been found in archaea or eucarya. Metal cations play an important role in cellular biochemistry. They are counter-ions of the negative charges in proteins, nucleic acids and phospholipids, and bridge these negative charges. Transitions metals serve as stability cores of proteins, catalyze redox reactions and rearrange small molecules. However, essential transition metal cations and those not needed by cells are toxic due to their interference with other metal cations, binding to wrong sites and dangerous redox reactions. It is therefore an interesting question how cells manage to allocate the correct metal to the right site and prevent others from doing harm. The first step into such a distribution process is a kinetic flow equilibrium of metal uptake and efflux processes that adjusts the cellular metal cation bouquet in just the right composition. Since metals may change their oxidation state or not but they cannot be degraded as organic molecules, metals are excellent subjects to understand basic homeostatic mechanisms of a living cell.
6. The Role of Efflux Pumps in the Nosocomial Pathogens Staphylococcus aureus and Acinetobacter baumannii
Bart A. Eijkelkamp, Karl A. Hassan, Ian T. Paulsen and Melissa H. Brown
Infections caused by opportunistic nosocomial pathogens can complicate the treatment of patients admitted to intensive care units. Most nosocomial bacterial pathogens possess an extended collection of resistance strategies to circumvent the effects of continuous exposure to the antimicrobial stresses present in these environments. Of these, active efflux has proven to be a successful detoxification mechanism employed by both Gram-positive and Gram-negative nosocomial pathogens, such as Staphylococcus aureus and Acinetobacter baumannii, respectively. Bioinformatic analyses of the genomes of these organisms determined that they each encode a large number of putative drug efflux systems including representatives from each of the five main families of efflux systems. Nonetheless, the repertoires of putative efflux systems encoded by these bacteria differ, primarily with respect to the dominant families of protective efflux systems; a trend that is likely to extend to most Gram-negative and Gram-positive bacteria. Various S. aureus transporters were among the first drug efflux systems to be described. Consequently, these proteins have been extensively characterized in work that has significantly advanced our fundamental knowledge of the structure and function of these complex proteins. In contrast, studies of A. baumannii efflux systems have been largely genetic, predominantly focused on explaining the high level of multidrug resistance observed in this bacterium without examining the biochemical or structural nature of the transporters themselves. Regulatory control of the genes encoding drug transporters is of major importance for multidrug resistance in both S. aureus and A. baumannii, since the overproduction of these proteins is typically detrimental to cell growth under non-selective conditions. Accordingly, it is not unusual for these systems to be controlled at a number of levels, both globally and locally by a range of regulatory factors. Here, we compare and contrast the efflux capabilities of these two bacterial pathogens.
7. Mycobacterium tuberculosis Drug Efflux Pumps: An Update
Maria Rosalia Pasca, Silvia Buroni and Giovanna Riccardi
It is well known that drug efflux systems contribute to the development of multi-resistance patterns in several bacterial pathogens. The selection and diffusion of Mycobacterium tuberculosis multidrug-resistant (MDR-TB), extensively drug-resistant (XDR-TB) and, more recently, totally drug-resistant (TDR) strains constitute a serious threat for tuberculosis global control. Mycobacteria, such as M. tuberculosis and Mycobacterium smegmatis, possess several putative drug efflux transporters, but their role in resistance is still a hard topic and needs to be further investigated as resistance to several drugs is usually the result of the combination of independent mutations in genes encoding either the drug target or the enzymes involved in drug activation. However, as the genetic basis of resistance to some antitubercular agents is not fully known for some clinical isolates, we cannot rule out an efflux mechanism in these strains. Several drug efflux transporters have been described in mycobacteria as responsible for resistance to aminoglycosides, chloramphenicol, fluoroquinolones, isoniazid, linezolid, rifampicin, tetracycline and other compounds but most of them were isolated in laboratory rather than in hospitals. This review highlights recent advances in our understanding of efflux-mediated drug resistance in mycobacteria, including the distribution of efflux systems in these organisms, their substrate profiles and their contribution to drug resistance.
8. Salmonella Efflux Pumps
Stephanie Baugh and Laura J. V. Piddock
Salmonella has around 350 efflux pumps which are important in the physiology of the bacterial cell. Nine of these pumps are known to confer low-level antibiotic resistance to three or more unrelated classes of drugs and so are known as multidrug resistant (MDR) efflux pumps. Salmonella possesses at least one MDR efflux pump from four of the five MDR transporter families; RND family, MATE family, MF superfamily and ABC superfamily. The MDR efflux system of most clinical importance, AcrAB-TolC, can export a wide variety of substrates. Antibiotic susceptibility testing of mutants in which specific genes have been inactivated reveals some overlapping substrate specificity of MDR efflux pumps. The efflux systems are under tight and well-ordered control from a variety of local and global regulatory genes, typically transcriptional activators such as RamA, although the precise nature of control is still to be elucidated. As well as being important in resistance of Salmonella to many antibiotics, dyes and detergents, MDR efflux pumps are also involved in virulence and biofilm formation revealing that they are fundamental to the biology of this important pathogen. Furthermore, these data suggest that AcrAB-TolC, or regulation thereof, could be targets for new inhibitors.
9. Pseudomonas aeruginosa Efflux Pumps
Antibiotic efflux systems are common in Pseudomonas aeruginosa, with chromosomally-encoded multidrug efflux systems of the Resistance Nodulation Division (RND) family, specifically MexAB-OprM, MexCD-OprJ, MexEF-OprN and MexXY-OprM, of particular importance in clinical settings. Despite the broad substrate specificity of many of these, their clinical importance is limited to fluoroquinolone resistance (MexAB-OprM, MexCD-OprJ and MexEF-OprN), β-lactam resistance (MexAB-OprM, MexXY-OprM) and aminoglycoside resistance (MexXY-OprM). Expression of these systems is governed by the products of regulatory genes (mexAB-oprM: mexR, nalC, nalD; mexCD-oprJ: nfxB; mexEF-oprN: mexT; mexXY: mexZ) whose mutation is typically responsible for acquired multidrug resistance in lab and clinical isolates. With few exceptions these efflux systems are not inducible by substrate antimicrobials, consistent with antimicrobial efflux not being their intended function. Indeed, recent data highlight their induction by environmental stresses (oxidative stress, nitrosative stress, envelope stress) suggestive of a role in stress response systems in this organism. Significantly, such stresses may provide a selective pressure for antibiotic-resistant efflux mutants in vivo independent of antibiotic exposure. Given the importance of these efflux systems in intrinsic and acquired multidrug resistance in P. aeruginosa, strategies aimed at interfering with efflux-mediated resistance are being investigated.
10. Function and Regulation of Neisseria gonorrhoeae Efflux Pumps
Yaramah M. Zalucki, Alexandra D. Mercante, Jason M. Cloward, Elizabeth A. Ohneck, Justin L. Kandler, Maira Goytia, Paul J.T. Johnson and William M. Shafer
The export action of efflux pumps is a nearly universal mechanism used by bacteria to escape the action of toxic compounds in their environment. Antimicrobials faced by bacteria include various biocides (natural or synthetic) and classical antibiotics used in therapy of infections. Certain efflux pumps also export antimicrobials produced by their hosts and this ability likely enhances the survival of the infecting pathogen, especially during early stages of infection when mediators of innate host defense normally function to reduce the microbial load. This review is concerned with the roles of efflux pumps produced by Neisseria gonorrhoeae in contributing to its resistance to antimicrobials used in therapy of infections or those that participate in innate host defense. Specific emphasis is placed on the genetic organization, transcriptional regulation, and function of gonococcal efflux pumps. The major theme of this review is that in addition to their role in enhancing bacterial resistance to classical antibiotics and biocides, certain efflux pumps, such as those harbored by strict human pathogens like gonococci, can also influence in vivo fitness and survival of bacteria since they provide a mechanism to resist natural antimicrobials produced by their host.
11. Multidrug Efflux Transporters in Campylobacter
Zhangqi Shen, Chih-Chia Su, Edward W. Yu and Qijing Zhang
As a major food-borne pathogen, Campylobacter is frequently exposed to antibiotics used for both animal production and human medicine. The increasing prevalence of antibiotic resistant Campylobacter has become a significant concern for public health. Among all known antibiotic resistance mechanisms, multidrug efflux systems play essential roles in the intrinsic and acquired resistance to structurally diverse antimicrobials. In Campylobacter, several multidrug efflux pumps, such as CmeABC, CmeDEF, CmeG, and Acr3, have been functionally characterized, which revealed that these efflux systems not only contribute to the resistance of antimicrobials, but also play important roles in facilitating the adaptation of Campylobacter to various environments, including the intestinal tract of animal hosts. The expression of these efflux transporters are controlled by transcriptional regulators, which sense the presence of toxic substrates and modulates the transcription of these efflux genes. Inhibiting the production or function of these multidrug efflux transporters, especially CmeABC, has been evaluated using efflux pump inhibitors and antisense peptide nucleic acid (PNA), demonstrating the potential of this approach for controlling antibiotic resistance in Campylobacter. In this Chapter, we will review the recent advance in understanding multidrug efflux systems and discuss the development of potential intervention strategies by targeting antimicrobial efflux pumps in Campylobacter.
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(EAN: 9781908230218 9781908230867 Subjects: [microbiology] [bacteriology] [medical microbiology] [molecular microbiology] [environmental microbiology] )