Efflux pumps are membrane transport proteins found in bacteria that pump antibiotics and other toxins out of bacterial cells, conferring resistance. There are five major classes of efflux pumps - SMR, MFS, RND, MATE, and ABC. RND pumps are the most prevalent in gram-negative bacteria and have three components. Efflux pump inhibitors can restore antibiotic susceptibility by competitively inhibiting the pumps. Efflux pumps play an important role in bacterial pathogenesis and survival by removing toxins from cells.

2. Antibiotics resistance via
Efflux pumps
Subhananthini jeyamurugan
18py17
II M.Sc., Microbiology

3. introduction
• Efflux pumps are elements that antibiotics are effective in resistance and are
involved in the pathogenesis of bacteria .
• Efflux pumps are important for processes of detoxification of Intracellular
metabolites, bacterial virulence in both animal an Man hosts, and etc…
• Efflux as a mechanism for antibiotic resistance was first described in 1980 .
• Multidrug efflux pumps cause serious problems in cancer chemotherapy and
the treatment of bacterial infections.

4. How does an efflux pump cause antibiotic
resistance?
• Drug efflux is a key mechanism of resistance in Gram-negative
bacteria.
• These systems pump solutes out of the cell.
• Efflux pumps allow the microorganisms to regulate their internal
environment by removing toxic substances, including antimicrobial
agents, metabolites and quorum sensing signal molecules.

5. Role of efflux pumps in antibiotic resistance
• Efflux pumps contribute to multidrug resistance as they expel
different types of antibiotics and chemicals such as dyes, organic
solvents, detergents, molecules needed for the cell–cell
communication, biocides, and metabolic products.

6. classification
• Efflux pump that was first detected : Efflux pumps was for a family of tetracycline
• Bacterial efflux transporters are classified into five major super families, based
on the amino acid sequence and the energy source used to export their
substrates.
• In the prokaryotic kingdom there are five major families of efflux transporters.

7. • The bacterial multidrug efflux transporters can be divided into five
classes:
(1) small multidrug resistance (SMR),
(2) major facilitator superfamily (MFS),
(3) resistance nodulation cell division (RND),
(4) multidrug and toxic compound extrusion (MATE),
(5) ATP-binding cassette (ABC).
Those five classes obtain energy required for the active transporting either from
H+ protons (RND, SMR, and MFS), Na+ dependent (MATE), or by hydrolysis ATP
(ABC).

8. • ABC: membrane proteins that translocate variety of substrates
across extra- and intra-cellular membranes ,
48 known ABC transporters; divided into 7 subfamilies of proteins
• RND: Fall into 7 phylogenetic families, 3 Restricted to Gram
negative Bacteria; other 4 either restricted to Gram positive or
represented in both also in bacteria, archaea and eukarya
• SMR: The smaller size . is unusual bacteria. Its pumps contain only
100-120 amino acid.

9. • MFS: One of two largest families, 17 MFS Families Present in
bacteria, archaea and eukarya
• MATE : 14 Phylogenetic Families, Also in bacteria, archaea and
eukarya . It has the same size MFS

10. classification
Schematic representation of the main types of bacterial
efflux systems

11. In gram positive
Bacteria.

12. In gram negative
Bacteria.

13. bacterial multidrug efflux transporters

14. Energy used in the efflux pump :
The energy source:
• ABC transporters are dependent on ATP hydrolysis;
(The primary active transport)
• MFS, RND and SMR are proton-driven efflux pumps (Proton motive force-driven and
ATP-dependent drug extrusion systems).
• MATE transporters consist of a Na/H drug anti porter system.
(Secondary active transport)

15. Gram-negative species with known efflux
systems:
• Haemophilus influenza
• Campylobacter jejuni
• Helicobacter pylori
• Vibrio parahaemolyticus
• Vibrio cholerae
• Neisseria gonorrhoeae...
• Salmonella Typhimurium
• Shigella dysenteriae
• Klebsiella pneumoniae
• Enterobacter aerogenes
• Serratia marcescens
• Proteus vulgaris
• Citrobacter freundii

16. Structure of drug efflux systems:
In Gram-positive bacteria:
A cytoplasmic membrane transport proteins that recognize
specific substrate. and the resistance.
In Gram-negative bacteria:
3 component in a system:
Two transport proteins within the membrane.
A periplasmic lipoprotein adapter

18. ABC transporters :
In contrast with prokaryotes, the major mechanism of efflux in eukaryotes is
dependent on proteins that derive their transport energy from the hydrolysis of
ATP.
Has a membrane-binding region of the cytoplasmic membrane is in addition to
the Alpha Helix to allow them The energy of ATP hydrolysis to transport the
drug, unlike other gradient used for this purpose are proton exchange
membrane of the electrochemical gradient.
Is an arsenic/antimony pump that is responsible for resistance to the antimonial
drug Pentostam in leishmania.

19. small multidrug resistance (SMR),
• The unusual bacteria found.
• One Of the 100 amino acids that made the four-helix
• For example :
EmrE is in E.coli
• QacH-2 resistance in Staphylococcus aureus causes a number of disinfectants.

21. The MFS (major facilitator superfamily) family :
• In eukaryotes, bacteria and Archaea are available.
• MFS are made of 400 amino acids arranged in 12 Helix membrane and a
cytoplasmic loop between helices 6 and 7 are large.
MFS transport of drugs in two classes be:
1- Class B: Transport tetracycline in E. Coli
2-class k: Transport tetracycline in Staphylococcus aureus

23. RND (resistance nodulation cell division) transporters :
• Typical emerged were larger than 1,000 amino acids. 12-helix structure. They
have a great second or subsequent intracytoplasmic Periplasmic between helices
1 and 2, 7 and 8.
• There are more gram-negative bacteria.
• Pump compounds: toxic metal ions, lipophilic drugs such as tetracycline,
quinolones, and beta-lactamase and chloramphenicol factors chemotherapy.

25. Structure :
DNA structure is composed of 3 components:
1-AcrA: inner membrane.
2-AcrB: inner membrane.
3-TolC: in the outer membrane.
The third monomer forming the inner membrane creates an area that
extends Periplasmic..

26. TOLC PROTEIN;
• Is a multifunctional protein. Moving in small drugs and toxins polypeptide
involved. for example: hemolysin .
• TolC is divided into two domains:
1-domain beta : in the outer membrane.
2-domain alpha : in the periplasmic space.
• TolC several actions have to cross the outer membrane periplasmic substrates.

28. •β-barrel :
Opens the outer membrane.
Beta consists of 12 strings and are arranged in a right-handed barrel.
•Α-helical :
12 left-handed filament is composed of two types of grape that are
Short
Tall

30. This substrate system:
Aminoglycosides
Tetracycline
Fluoro quinolones
Chloramphenicol
Trimethoprim

32. MATE transporters
• Of 450 amino acids that have been arranged in 12 Helix These trans Porter Na or
protons act as anti ports.
• NorM resistance in Vibrio parahaemolyticus color, aminoglycosides and
fluoroquinolones cause.
• YdhE in E.Coli is a cationic resistance to antibiotics cause
• MepA in the transfer tetracycline Staphylococcus aureus, detergents and paints
as well.
• PepM in substrate fluoroquinolones, is ethidium bromide and disinfectants

33. The pump substrates include:
•Gentamicin
• Ciprofloxacin
• Erythromycin
• Trimethoprim

34. What are efflux pump inhibitors?
• Peptidomimetic compounds such as phenylalanine arginyl β-
naphthylamide (PAβN) have been introduced as efflux pump
inhibitors (EPIs).
• Their mechanism of action is through competitive inhibition with
antibiotics on the efflux pump resulting in increased intracellular
concentration of antibiotics.

35. Inhibitors of efflux pumps have great potential as pharmacological agents that
restore the drug susceptibility of multidrug resistant bacterial pathogens.

36. • High-potential pharmacological agents that can be multi-
drug resistant pathogens return sensitivity to the drug.
• Example:
1) Phenylalanine, arginine beta - naphthyl amide
2) verapamil
3) naphthyl piperidine
4) chlorpromazine

37. Phenyl-Arginyl ß N-naphtylamide
verapamil

38. naphthyl piperidine
chlorpromazine

39. The importance of efflux pumps:
• Promotes the excretion of drugs and chemicals are.
• Metabolites and toxins that repel bacteria
• Microorganisms against antibiotics and chemicals, toxins, stress and protect
• The survival of microorganisms in different environments.
• The supply of materials for the synthesis of bacterial surface structures involved
• In balancing solutions, ionic homeostasis and balance are important.