Cyclic lipopeptides, folate antagonists, licosamide, polymixins

1. PHAR 532
LECTURE 9
CYCLIC LIPOPEPTIDES, FOLATE
ANTAGONISTS, LICOSAMIDE,
POLYMIXINS
Antimicrobial agents and
Immunization

2. Cationic antimicrobial peptides
Despite the diversity in the amino acid sequence and
the structural classes (i.e., β-sheets, α-helices, loops
and extended structures) of antimicrobial peptides,
with over 700 known to date, they all share a
common three-dimensional arrangement.
They fold into amphiphilic molecules, with one face
being hydrophobic, while the other is charged

3. Fig. 1. Model for the mechanism of action of cationic antimicrobial peptides. The unstructured peptides (a) adopt secondary
structure upon interaction with the bacterial membrane, and particularly anionic phospholipids. (b) This structure can consist of β-
shee...
Suzana K. Straus, Robert E.W. Hancock
Mode of action of the new antibiotic for Gram-positive pathogens daptomycin: Comparison with cationic antimicrobial
peptides and lipopeptides
Biochimica et Biophysica Acta (BBA) - Biomembranes, Volume 1758, Issue 9, 2006, 1215–1223
http://dx.doi.org/10.1016/j.bbamem.2006.02.009

4. Lipopeptides
Lipopeptides consist of a linear or cyclic peptide
sequence, with either net positive or negative charge,
to which a fatty acid moiety is covalently attached to
its N-terminus.
Highly active against multi-resistant bacteria. Some
lipopeptides also display anti-fungal activity.

5. Lipopeptides
In the anionic lipopeptide class, the first naturally
occurring member to be discovered was
amphomycin over fifty years ago.
Additional members of this class of compounds
include crystallomycin, aspartocin, glumamycin,
laspartomycin, tsushimycin, and daptomycin.

6. Lipopeptides mode of action
They are believed to target and bind to the bacterial
membrane directly and cause rapid depolarization of
the antibacterial membrane potential.

7. Daptomycin

8. Daptomycin
Naturally occurring compound found in the
soil saprotroph Streptomyces roseosporus.
The antibacterial activity of daptomycin is strictly
dependent on the presence of physiological levels of
free calcium ions.
Daptomycin disrupts Gram-positive cytoplasmic
membrane function, causing leakage of potassium
(and potentially other) ions, ultimately leading to
loss of membrane potential and cell death.

9. Daptomycin mechanism of action.
Judith N. Steenbergen et al. J. Antimicrob. Chemother.
2005;55:283-288
JAC vol.55 no.3 © The British Society for Antimicrobial Chemotherapy 2005; all rights reserved

10. Resistance
Many of the mutations that alter susceptibility to
daptomycin have been shown to directly affect
membrane lipid composition.
MprF is a membrane protein responsible for
synthesizing a positively charged phospholipid,
lysylphosphatidylglycerol. In S. aureus, mutations
that eliminate the expression of the mprF gene
sensitize cells to daptomycin, and mutations that
confer resistance are believed to be “gain-of-
function” alleles that increase the amount of
lysylphosphatidylglycerol.

11. Resistance
The specific membrane modifications that confer
resistance vary by species: in enterococci, changes in
cardiolipin synthesis are associated with resistance,
while in Bacillus subtilis, resistance may be linked to
the overall phosphatidylglycerol content due to
mutations in pgsA

12. Folate antagonists – Sulfonamides
Sulfonamides inhibit the synthesis of folic acid by
competing with PABA for the active site of an
enzyme involved in the pathway.
Sulfonamides are bacteriostatic against S. aureus.
Some strains of S. aureus overproduce para-
aminobenzoic acid (PABA), causing resistance to
sulfonamides. However, resistance to sulfonamides
is more frequently caused by alterations in
dihydropteroate synthase.

13. Folate antagonists – Trimethoprim
Trimethoprim is a tetrahydrofolate reductase
inhibitor that, when added to sulfamethoxazole,
provides a second step block in the folate
biosynthetic pathway.

14. Sulfonamide and trimethoprim

15. Folate antagonists and the biosynthesis of thymidine.
Ellie J. C. Goldstein, and Richard A. Proctor Clin Infect
Dis. 2008;46:584-593
© 2008 Infectious Diseases Society of America

16. Resistance
Mutations in the gene for dihydropteroate synthase
decrease affinity for sulfonamides.
Mutations in the chromosomal gene specifying
dihydrofolate reductase can result in enzyme with
reduced affinity for trimethoprim.
Acquired resistance through plasmids and integrons
is also possible.
sul1 and sul2 are the main determinants of clinical
resistance to sulfonamide.
dfr1-20 confer resistance to trimethoprim.

17. Lincosamides
Lincosamides are a class of antibiotics which
include lincomycin, clindamycin, and pirlimycin.
Lincosamides prevent bacteria replicating
by interfering with the synthesis of proteins. They
bind to the 23s portion of the 50S subunit of
bacterial ribosomes and cause premature
dissociation of the peptidyl-tRNA from the ribosome.

18. Lincomycin

19. Clyndamycin

20. Resistance
Methylation of the ribosome target. Ribosomal
methylation is catalyzed by members of the
erythromycin ribosome methylase (erm) enzymes.

21. Polymixins
Polymyxins are a group of polypeptide antibiotics
that consists of five chemically different compounds
(polymyxins A–E).
General structure: cyclic peptide with a
long hydrophobic tail.
Only polymyxin B and polymyxin E (colistin) have
been used in clinical practice. They differ by a single
amino acid change (D-phenylalanine in polymixin B
replaces D-leucine in colistin). The mechanism
behind colistin's bactericidal ability is considered to
be identical to that of polymyxin.

22. Polymixin B

23. Colistin

24. Polymixins
The initial target of the antimicrobial activity of
polymyxins is the lipopolysaccharide (LPS)
component of the outer membrane.

25. Molecular models of the
complex between
polymyxin B1 and the lipid
A structure from Klebsiella
pneumoniae
(A)The lipid A molecule is
shown in space-filling
representation and polymyxin
B1 is shown in stick
representation.
(B) The chemical structure of
the lipid A molecule.
(C) The key electrostatic
interactions between
positively charged Dab
residues on polymyxin and
the negatively charged lipid A
phosphoresters.
Future Microbiol. 2013 Jun;

26. Resistance
It is becoming increasingly apparent that polymyxin
resistance in Gram-negative bacteria involves the
multitier upregulation of a number of regulatory
systems. LPS remodeling is an important survival
strategy for Gram-negative bacteria.
The most common polymyxin resistance mechanism
in P. aeruginosa, Salmonella enterica serovar
Typhimurium, E. coli, A. baumannii and K.
pneumoniae is due to modifications of lipid A
phosphates with positively charged groups.

27. Resistance
A number of unique and often species-specific polymyxin
resistance mechanisms do not involve the LPS-binding
pathway.
Capsular polysaccharide levels on K. pneumoniae have
been shown to coincide with polymyxin resistance.
Polymyxin resistance in a number of Gram-negative
bacterial species has been associated with alterations in
the expression of outer membrane proteins, including
efflux pumps. In P. aeruginosa biofilms, colistin
resistance was found to coincide with the overexpression
of the mexAB–oprM efflux pump system.

28. Resistance
Polymyxin resistance in P. aeruginosa has also been
associated with changes in the expression of the outer
membrane protein OprH, which is purported to perform a
membrane stabilization role under conditions of
Mg2+
starvation.
In K. pneumoniae, a deficiency in the outer membrane
protein OmpA, which mediates adhesion to eukaryotic cells,
has been associated with an increased susceptibility to
polymyxin B.
Moreover, the AcrAB–TolC energy-driven efflux pump has
been linked to polymyxin resistance and efflux from K.
pneumoniae and E. coli cells. In Burkholderia vietnamien, a
multidrug efflux pump NorM has been shown to contribute to
polymyxin resistance.

29. Polymixin resistance
Future Microbiol. 2013 Jun; 8(6)