Probing Nature for Antibiotics

1
Probing Nature for Antibiotics

• Irosha Nayanthika Nawarathne
• Michigan State University
• 04/30/08

health.howstuffworks.com
2
Struggle for living
dansaper.blogspot.com, www.photos-screensaver-make
r.com, tecnocientista.info.com,
www.creswell-crags.org.uk
3

• History of humankind can be regarded from a
  medicinal point of view as a struggle against
  infectious diseases

Yoneyama, H., Katsumata, R., Biosci. Biotechnol.
Biochem., 2006, 70,1060
4
Survival against infectious diseases
dodd.cmcvellore.ac.in, www.ayurvedicmedicine4u.com
, www.rootsweb.com
5
What are antibiotics?
Molecules that stop the microbial growth (both
bacteria and fungi) or kill them outright
Walsh, C., Antibiotics Actions origins and
Resistance, 2003, 4
6
How do the antibiotics act against bacteria?
Cell Wall Biosynthesis ß-lactams,Cyclospori
ns,Glycopeptides
Walsh, C., Antibiotics Actions origins and
Resistance, 2003, 19 www.jacksofscience.com
7
How do the antibiotics act against bacteria?
Protein Biosynthesis Aminoglycosides,Macrolides, T
etracyclines,Oxazolidinones
Walsh, C., Antibiotics Actions origins and
Resistance, 2003, 19 www.istockphoto.com
8
How do the antibiotics act against bacteria?
DNA Biosynthesis Quinolones
RNA Biosynthesis Rifampicin
Walsh, C., Antibiotics Actions origins and
Resistance, 2003, 19 publications.nigms.nih.gov,
www.istockphoto.com
9
How do the antibiotics act against bacteria?
Metabolic pathways Folic Acid
Metabolism Trimethoprim, Sulfonamides Fatty
Acid Biosynthesis Triclosan, Isoniazid,
Ethionamide
Walsh, C., Antibiotics Actions origins and
Resistance, 2003, 19 www.istockphoto.com
10
Why do we need more antibiotics?

• - Developing antimicrobial resistance
• Bacterial species Common types of Antimicrobial
  Types of Infections
  
  
  Resistance
• Streptococcus pneumoniae ß-lactams,
  cephalosporins, macrolides Otitis media,
  pneumonia,
• Tetracyclines sinusitis, meningitis
• Staphylococcus aureus
• Community-associated Meticillin, cephalosporins,
  macrolides Skin, soft tissue, sepsis
• pneumonia
• Healthcare-associated Meticillin,
  cephalosporins, quinolones, Endocarditis,
  pneumonia,
• aminoglycosides, macrolides sepsis
• Enterococcus spp. Ampicillin, vancomycin,
  aminoglycosides Sepsis, urinary tract

Furuya, E.Y., Lowy, F.D., Nature, 2006, 4, 36
11
What should be targeted?

• The compounds with,
• Novel structures
• New modes of action

Fernandes, P., Nature Biotechnology, 2006, 24,
1497
12

• Where do the antibiotics come from?

NATURE
13

• Where do the antibiotics come from?

NATURE
TS
NP
SS
14

• Where do the antibiotics come from?

NATURE
Helps in designing the molecules
TS
NP
SS
15
Natural products as antibiotics

• Naturally occurring compounds that are end
  products of secondary metabolism.
• Mostly extracted from plants, marine organisms,
  or microorganisms.

Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006
16
Natural products as antibiotics

• Naturally occurring compounds that are end
  products of secondary metabolism.
• Mostly extracted from plants, marine organisms,
  or microorganisms.
• Eg

Isolation - Streptomyces erythreus in 1952 Uses
- Respiratory tract diseases,
genital infections MOA - Inhibition of
protein synthesis
Erythromycin
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 Pal, S.,
Tetrahedron, 2006, 62, 3171
17
Antibiotics which are semi-synthesized

• Synthetically modified chemical compounds which
  are originated
• from natural products.

Walsh, C., Antibiotics Actions origins and
Resistance, 2003, 4
18
Erythromycin is
Acid unstable
Pal, S., Tetrahedron, 2006, 62, 3171
19
Antibiotics which are semi-synthesized
Clarithromycin
Azithromycin
HMR3647
TE802
Pal, S., Tetrahedron, 2006, 62, 3171
20
Antibiotics which are totally from synthesis

• Totally synthesized molecules which are potent as
  antibiotics.
• Three main types.
• 1. Sulfa drugs
• 2. Quinolones
• 3. Oxazolidinones

Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006
21
Antibiotics which are totally from synthesis

• Sulfa drugs (Sulphonamides)

Sulfamethoxazole
Uses - Urinary tract infections, pneumonia
etc. MOA - Inhibition of folate synthesis

Harold, P.L., OGrady, F.W., Antibiotic and
Chemotherapy, 1992, 6, 268-272
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006
22
Antibiotics which are totally from synthesis

• Sulfa drugs (Sulphonamides) Naturally
  occurring

Sulfamethoxazole
p-aminobenzoic acid
Uses - Urinary tract infections, pneumonia
etc. MOA - Inhibition of folic acid
biosynthesis
Harold, P.L., OGrady, F.W., Antibiotic and
Chemotherapy, 1992, 6, 268-272
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 Walsh, C.,
Antibiotics Actions origins and Resistance, 2003,
80-82
23
Antibiotics which are totally from synthesis

• Quinolones

Ciprofloxacin
Uses - Urinary tract infections, Lower
respiratory infections, Gastrointestinal
infections MOA - Inhibition of
DNA synthesis
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006
24
Antibiotics which are totally from synthesis

• Quinolones
  Naturally occurring

Aurachin D
Ciprofloxacin
Aurachin C
Uses - Urinary tract infections, Lower
respiratory infections, Gastrointestinal
infections MOA - Inhibition of
DNA synthesis
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 Kunze, B., Hofle,
G., Reichenbach, H., J. Antibiotics, 1987, 40, 258
25
Antibiotics which are totally from synthesis

• Oxazolidinones

Linezolid
Uses - Soft tissue infections, skin
infections, Tuberculosis etc. MOA -
Inhibition of protein synthesis
Ford, C.W., Zurenko, G.E., Barbachyn, M.R.,
Current Drug Targets-Infectious Disorders, 2001,
1,181
26
Antibiotics which are totally from synthesis

• Oxazolidinones
  Naturally occurring

(-)-Cytoxazone
Linezolid
()-Sreptazolin
Uses - Soft tissue infections, skin
infections, Tuberculosis etc. MOA -
Inhibition of protein synthesis
Ford, C.W., Zurenko, G.E., Barbachyn, M.R.,
Current Drug Targets-Infectious Disorders, 2001,
1,181 Zappia, G., et al., Mini-Reviews in
Medicinal Chemistry, 2007, 7, 389
27
Sources of antibacterial drugs from 1981 to 2002
Newman, D.J., Cragg, G.M., Snader, K.M., J. Nat.
Prod., 2003, 66, 1022
28

• Ways of probing nature for antibiotics

29

• Ways of probing nature for antibiotics

New antibiotics New architectural scaffolds
30
Approach A

• Conventional way of NP discovery

Extraction to the solvents
Natural materials
Isolation and Structure Elucidation
Bioassay guided fractionation
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 www.spc.int,
www.oceanexplorer.noaa.gov, www.nature.com,
www.textbookofbacteriology.net
31
Approach A

• Conventional way of NP discovery
• Why isnt it successful?
• Problems associated with the growth or the
  availability of the source
• Replication of the hits
• Do not distinguish novel from old
• Mostly miss the novel compounds due to the lack
  of sensitivity
• No hints about MOA
• Cannot reveal potency at screening stage

Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 Clardy, J.,
Fischbach, M.A., Walsh, C., Nat. Rev.
Biotechnol., 2006, 24, 1541
32
Approach A

• What are the new strategies to explore nature for
  NPs

Novel culturing techniques
Heterologous expression of biosynthetic genes
Metagenomics
Molecular Biology based Techniques
Genomics and Combinatorial biosynthesis
Precursor directed biosynthesis Mutasynthesis
Differential sensitivity screening approach
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 Clardy, J.,
Fischbach, M.A., Walsh, C., Nat. Rev.
Biotechnol., 2006, 24, 1541 Donadio, S.,
Chemistry Biology, 2006, 13, 560
33
Approach A

• What are the new strategies to explore nature for
  NPs

Novel culturing techniques
Heterologous expression of biosynthetic genes
Metagenomics
Molecular Biology based Techniques
Genomics and Combinatorial biosynthesis
Precursor directed biosynthesis Mutasynthesis
Differential sensitivity screening approach
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 Clardy, J.,
Fischbach, M.A., Walsh, C., Nat. Rev.
Biotechnol., 2006, 24, 1541 Donadio, S.,
Chemistry Biology, 2006, 13, 560
34
Approach A

• Precursor Directed Biosynthesis Mutasynthesis

Extraction to the Solvents
Producing organisms found in nature
Pathogen
Wild type
Mutant type
35
Approach A

• Precursor Directed Biosynthesis Mutasynthesis

Wild type
Natural Biosynthetic pathway
Kennedy, J., Nat. Prod. Rep., 2008, 25, 25 Weist,
S., S?ssmuth, R. D., Appl. Microbiol.
Biotechnol., 2005, 68, 141
36
Approach A

• Precursor Directed Biosynthesis and Mutasynthesis

Wild type
Precursor-Directed Biosynthesis
Kennedy, J., Nat. Prod. Rep., 2008, 25, 25 Weist,
S., S?ssmuth, R. D., Appl. Microbiol.
Biotechnol., 2005, 68, 141
37
Approach A

• Precursor Directed Biosynthesis and Mutasynthesis

Mutant
Mutasynthon
Mutasynthesis
Kennedy, J., Nat. Prod. Rep., 2008, 25, 25 Weist,
S., S?ssmuth, R. D., Appl. Microbiol.
Biotechnol., 2005, 68, 141
38
Approach A

• Mutasynthesis

Ring A Ring B Ring C Novobiocin
(Albamycin)
Ring A Ring B Ring C Clorobiocin
Pojer, F., Li, S.M., Heide, L., Microbiology,
2002, 148, 3901 Galm, U., et al, Chemistry
Biology, 2004, 11, 173 Weist, S., S?ssmuth, R.
D., Appl. Microbiol. Biotechnol., 2005, 68, 141
39
Approach A

• Mutasynthesis

CloQ- mutants
Pojer, F., Li, S.M., Heide, L., Microbiology,
2002, 148, 3901 Galm, U., et al, Chemistry
Biology, 2004, 11, 173 Eust?quio, A.S., et al,
Arch. Microbiol., 2003, 180, 25
40
Approach A

• Mutasynthesis

Clorobiocin
CloQ-mutant
Galm, U., et al, Chemistry Biology, 2004, 11,
173 Pojer, F., Li, S.M., Heide, L.,
Microbiology, 2002, 148, 3901
41
Approach A

• Mutasynthesis

Clorobiocin
CloQ-mutant
Galm, U., et al, Chemistry Biology, 2004, 11,
173 Pojer, F., Li, S.M., Heide, L.,
Microbiology, 2002, 148, 3901
42
Approach A

• Mutasynthesis

CloQ-mutant
Analogs of Clorobiocin
Galm, U., et al, Chemistry Biology, 2004, 11,
173 Galm, U., et al, Antimicrob. Agents
Chemother., 2004, 48, 1307 Pojer, F., Li, S.M.,
Heide, L., Microbiology, 2002, 148, 3901
43
Approach A

• What are the new strategies to explore nature for
  NPs

Novel culturing techniques
Heterologous expression of biosynthetic genes
Metagenomics
Molecular Biology based Techniques
Genomics and Combinatorial biosynthesis
Precursor directed biosynthesis Mutasynthesis
Differential sensitivity screening approach
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 Clardy, J.,
Fischbach, M.A., Walsh, C., Nat. Rev.
Biotechnol., 2006, 24, 1541 Donadio, S.,
Chemistry Biology, 2006, 13, 560
44
Approach A

• Differential sensitivity screening approach

Producing organism from nature
Pathogen
Expression of certain protein/s
Wild type
Normal
Extraction to the solvents
Low
Disabled type
Increased sensitivity
Target the pathway
Couzin, J., Nature, 2006, 314, 34, Forsyth R.A.,
Molecular Biology, 2002, 43, 1387 Wang, J., et
al, Antimicrob. Agents Chemother., 2006, 50, 519
45
Approach A

• Differential sensitivity screening approach

Fatty Acid Biosynthesis A good target
FAB Type I - In mammals
FAB Type II - In bacteria
Campbell, J.W., Cronan, J.E.Jr., Annu. Rev.
Microbiol., 2001, 55, 305
46
Biosynthesis of Saturated Fatty Acids
Campbell, J.W., Cronan, J.E.Jr., Annu. Rev.
Microbiol., 2001, 55, 305
47
Biosynthesis of Saturated Fatty Acids
Campbell, J.W., Cronan, J.E.Jr., Annu. Rev.
Microbiol., 2001, 55, 305
48
Biosynthesis of Saturated Fatty Acids
Campbell, J.W., Cronan, J.E.Jr., Annu. Rev.
Microbiol., 2001, 55, 305
49
Approach A

• Differential sensitivity screening approach

RNA-mediated gene silencing technique
Singh, S.B., et al, J. Am. Chem. Soc., 2006, 128,
11916 Forsyth, R.A., Molecular Biology, 2002, 43,
1387 Wang, J., et al, Antimicrob.
Agents Chemother., 2006, 50, 519
50
Approach A
Differential sensitivity screening approach
RNA-mediated gene silencing technique
In Prokaryotes-
ds RNA
5 AUGGCCUGGACUUCA3 3 UACCGGACCTGTTGU
5
Degradation of fabF mRNA or inhibition of
translation
Reduced or No FabF expression
Higher sensitivity towards FabF inhibitors
Singh, S.B., et al, J. Am. Chem. Soc., 2006, 128,
11916 Forsyth, R.A., Molecular Biology, 2002, 43,
1387 Wang, J., et al, Antimicrob.
Agents Chemother., 2006, 50, 519
51
Approach A

• Differential sensitivity screening approach

Results - RNA-mediated gene silencing technique
Wild type fabF Anti-sense
Inhibitor (µg)
Wang, J., et al, Nature, 2006, 441, 358
52
Approach A

• Differential sensitivity screening approach

Results - RNA-mediated gene silencing technique
Wild type fabF Anti-sense
200 times more potent than Cerulenin
Wild type fabF Anti-sense
Inhibitor (µg)
Wang, J., et al, Nature, 2006, 441, 358 Price,
A.C., et al, The Journal of Biological Chemistry,
2001, 276, 6551 Heath, R.J., White, S.W., Rock,
C.O., Progress in Lipid Research, 2001, 40, 467
53
Approach A

• Differential sensitivity screening approach

Discovery of Platensimycin
Platensimycin from a strain of Streptomyces
platensis
Singh, S.B., et al, J. Am. Chem. Soc., 2006, 128,
11916
54
Approach A

• Differential sensitivity screening approach

Potency of Platensimycin
Organism and genotype Platensimycin
Linezolid Antibacterial activity
(MIC, µg/ml)
S. aureus (MSSA) 0.5 4 S. aureus (MRSA)
0.5 2 S. aureus (MRSA, macrolideR) 0.5
2 S. aureus (MRSA, linezolidR) 1
32 Enterococcus faecium (VRE)
0.1 2
Toxicity
(µg/ml) HeLa MTT (IC50)
gt1,000 gt100
MIC Concentration of inhibitor used to result
no visible growth of the pathogens
IC50 Concentration of the inhibitor used to
kill 50 population of the living cells
Wang, J., et al, Nature, 2006, 441, 358
55
Approach A
Differential sensitivity screening approach
High FabF selectivity

• Cell - free gel - elongation assay

Malonyl-ACP
C41(?2)-ACP
C40-ACP
gt6C-ACP
Wang, J., et al, Nature, 2006, 441, 358
Heath, R.J., Nat.Prod.Rep., 2002, 19, 581
56

• Ways of probing nature for antibiotics

New antibiotics New architectural scaffolds
57
Approach B
Generating Nature Mimics
Biosynthetic pathway
Enzyme purification 3D structural determination
Designing theoretical chemical space that fits
the active site or docking the database
structures
Translate to a real structure by synthesis
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 Clardy, J.,
Fischbach, M.A., Walsh, C., Nat. Rev.
Biotechnol., 2006, 24, 1541
58
Approach B
Generating Nature Mimics
Biosynthesis of lysine A good target

• Essential for the bacterial growth
• Does not exist in mammals

Hutton, C.A., Perugini, M.A., Gerrard, J.A., Mol.
Biosyst., 2007, 3, 458 Hutton, C.A., Southwood,
T.J., Turner, J.J., Mini-Reviews in Medicinal
Chemistry, 2003, 3,115
59
Biosynthesis of lysine
methionine
threonine
isoleucine
Hutton, C.A., Perugini, M.A., Gerrard, J.A., Mol.
Biosyst., 2007, 3, 458 Hutton, C.A., Southwood,
T.J., Turner, J.J., Mini-Reviews in Medicinal
Chemistry, 2003, 3,115
60
Biosynthesis of lysine
Hutton, C.A., Perugini, M.A., Gerrard, J.A., Mol.
Biosyst., 2007, 3, 458 Hutton, C.A., Southwood,
T.J., Turner, J.J., Mini-Reviews in Medicinal
Chemistry, 2003, 3,115
61
Approach B
Generating Nature Mimics
Proposed mechanism
Hutton, C.A., Perugini, M.A., Gerrard, J.A., Mol.
Biosyst., 2007, 3, 458 Hutton, C.A., Southwood,
T.J., Turner, J.J., Mini-Reviews in Medicinal
Chemistry, 2003, 3,115
62
Approach B
Generating Nature Mimics
Supportive data
Faehnle, C.R., Coq, J.L., Liu, X., Viola, R.E.,
Journal of Biological Chemistry, 2006, 281, 31031
Cox, R.J., Gibson, J.S., Martín, M.B.M.,
ChemBioChem, 2002, 3, 874 Hutton, C.A.,
Southwood, T.J., Turner, J.J., Mini-Reviews in
Medicinal Chemistry, 2003, 3,115
63
Approach B
Generating Nature Mimics
Inhibitors of lysine biosynthesis
Cox, R.J., Gibson, J.S., Martín, M.B.M., Chem.
Commun., 2001, 1710 Cox, R.J., Gibson, J.S.,
Hadfield, A.T., ChemBioChem, 2005, 6, 2255 Cox,
R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem,
2002, 3, 874
64
Approach B
Generating Nature Mimics
Inhibitors of lysine biosynthesis
Cox, R.J., Gibson, J.S., Martín, M.B.M., Chem.
Commun., 2001, 1710 Cox, R.J., Gibson, J.S.,
Hadfield, A.T., ChemBioChem, 2005, 6, 2255 Cox,
R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem,
2002, 3, 874
65
Approach B
Generating Nature Mimics
In vitro assays
Reverse Biosynthesis
Cox, R.J., Gibson, J.S., Martín, M.B.M.,
ChemBioChem, 2002, 3, 874 Cox, R.J., Gibson,
J.S., Hadfield, A.T., ChemBioChem, 2005, 6, 2255
66
Approach B
Generating Nature Mimics
Competitive assays
Direct assay
KI (ASA) KI (Phosphate)
- -
750 µM 2130µM
214 µM 92µM
Cox, R.J., Gibson, J.S., Martín, M.B.M.,
ChemBioChem, 2002, 3, 874 Blanco, J., Moore,
R.A., Viola, R.E., PNAS, 2003, 100, 12613 Han,
S., Moore, R.A., Viola, R.E., Synlett, 2003, 6,
845
67
Approach B
Generating Nature Mimics
Competitive assays
Direct assay
Cox, R.J., Gibson, J.S., Martín, M.B.M.,
ChemBioChem, 2002, 3, 874 Blanco, J., Moore,
R.A., Viola, R.E., PNAS, 2003, 100, 12613 Han,
S., Moore, R.A., Viola, R.E., Synlett, 2003, 6,
845
68
Approach B
Generating Nature Mimics
Competitive assays
Direct assay
Cox, R.J., Gibson, J.S., Martín, M.B.M.,
ChemBioChem, 2002, 3, 874 Blanco, J., Moore,
R.A., Viola, R.E., PNAS, 2003, 100, 12613 Han,
S., Moore, R.A., Viola, R.E., Synlett, 2003, 6,
845
69
Approach B
Generating Nature Mimics
Competitive assays
Direct assay
KI (ASA) KI (Phosphate) 2nd pKa
- - 4.2-5.0
750 µM 2130µM 6.1
214 µM 92µM 6.2-6.4
Cox, R.J., Gibson, J.S., Martín, M.B.M.,
ChemBioChem, 2002, 3, 874 Blanco, J., Moore,
R.A., Viola, R.E., PNAS, 2003, 100, 12613 Han,
S., Moore, R.A., Viola, R.E., Synlett, 2003, 6,
845
70
Approach B
Generating Nature Mimics
Time-dependent inhibition assays
Pre-incubation assay
KI (ASA)
95µM
-
-
Cox, R.J., Gibson, J.S., Martín, M.B.M.,
ChemBioChem, 2002, 3, 874 Blanco, J., Moore,
R.A., Viola, R.E., PNAS, 2003, 100, 12613 Han,
S., Moore, R.A., Viola, R.E., Synlett, 2003, 6,
845
71
Approach B
Generating Nature Mimics
Time-dependent inhibition assays
Pre-incubation assay
Cox, R.J., Gibson, J.S., Martín, M.B.M.,
ChemBioChem, 2002, 3, 874 Blanco, J., Moore,
R.A., Viola, R.E., PNAS, 2003, 100, 12613 Han,
S., Moore, R.A., Viola, R.E., Synlett, 2003, 6,
845
72

• Ways of probing nature for antibiotics

New antibiotics New architectural scaffolds
73
Please, Dont flush!

• Average american receives more than 11
  prescriptions a year.
• About 3.3 billion a total.
• Nonprescription drugs !

Halford, B., C EN News, 2008, 86, 13 Halford,
B., C EN News, 2008, 86, 16
74
AcknowledgementDr. WalkerDr. HausingerDr.
ArnostiDr. StoltzfusDr. Stephen Soisson, Dr.
Jun Wang (Merck)Labmates - Behnaz, Danielle,
Joshua, Mark, Washington, Yemane


Friends -
Samantha, Sue, Tharanga, Xiaofei
75
Thank you all !
76
Back-up slides


77
Approach A

• Differential sensitivity screening approach

In vivo studies of Platensimycin
In a mouse model of disseminated S. aureus
infection
Wang, J., et al, Nature, 2006, 441, 358
78
Timeline of discovery of novel classes of
antibiotics and introduction in clinic
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006
79
Approach B
Generating the Nature Mimics
Faehnle, C.R., Coq, J.L., Liu, X., Viola, R.E.,
Journal of Biological Chemistry, 2006, 281, 31031
Hutton, C.A., Perugini, M.A., Gerrard, J.A.,
Mol. Biosyst., 2007, 3, 458 Hutton, C.A.,
Southwood, T.J., Turner, J.J., Mini-Reviews in
Medicinal Chemistry, 2003, 3,115
80
Antibiotics which are totally from synthesis

• Sulfa drugs (Sulphonamides) Naturally
  occurring

Sulfamethoxazole
p-aminobenzoic acid
Uses - Urinary tract infections, pneumonia
etc. MOA - Inhibition of folate synthesis

Harold, P.L., OGrady, F.W., Antibiotic and
Chemotherapy, 1992, 6, 268-272
Singh, S.B., Barrett, J.F., Biochemical
Pharmacology, 2006, 71, 1006 Walsh, C.,
Antibiotics Actions origins and Resistance, 2003,
80-82
81
Approach A

• Precursor Directed Biosynthesis

Penicillium Chrysogenum (Penicillium notatum)
6-APA
Nayer, J.H.C., Trends. Biochem. Sci., 1991, 16,
195 Nayer, J.H.C., Trends. Biochem.Sci., 1991,
16, 234 Kennedy, J., Nat. Prod. Rep., 2008, 25,
25
82
Approach A

• Precursor Directed Biosynthesis

Penicillin G
Penicillium Chrysogenum (Penicillium notatum)
Penicillin V
Nayer, J.H.C., Trends. Biochem. Sci., 1991, 16,
195 Nayer, J.H.C., Trends. Biochem.Sci., 1991,
16, 234 Kennedy, J., Nat. Prod. Rep., 2008, 25,
25
83
Approach A

• Mutasynthesis

Nov L
Clo L
Ring A Ring B Ring C Novobiocin
(Albamycin)
Ring A Ring B Ring C Clorobiocin
Pojer, F., Li, S.M., Heide, L., Microbiology,
2002, 148, 3901 Galm, U., et al, Chemistry
Biology, 2004, 11, 173 Eust?quio, A.S., et al,
Arch. Microbiol., 2003, 180, 25
84

• Where do the antibiotics come from?

NATURE
Kekule stucture of benzene
TS
NP
SS
www.boomeria.org
85
Approach A

• Precursor Directed Biosynthesis
• Drawbacks
• Involves complex purification procedures
• Require high concentrations of synthetic
  precursor
• Only few intermediates will incorporate into the
  product

Kennedy, J., Nat. Prod. Rep., 2008, 25, 25 Weist,
S., S?ssmuth, R. D., Appl. Microbiol.
Biotechnol., 2005, 68, 141
86
Inhibitors of Fatty Acids Biosynthesis
Triclosan, Isoniazid, Ethionamide
Continues...
Cerulenin, Thiolactomycin
Campbell, J.W., Cronan, J.E.Jr.,
Annu.Rev.Microbiol., 2001, 55, 305 Price, A.C.,
et al, The Journal of Biological Chemistry, 2001,
276, 6551 Heath, R.J., White, S.W., Rock, C.O.,
Progress in Lipid Research, 2001, 40, 467
87

• Where do the antibiotics come from?

NATURE
88
Approach B
Generating the Nature Mimics
Cox, R.J., Gibson, J.S., Martín, M.B.M., Chem.
Commun., 2001, 1710 Cox, R.J., Gibson, J.S.,
Hadfield, A.T., ChemBioChem, 2005, 6, 2255 Cox,
R.J., Gibson, J.S., Martín, M.B.M., ChemBioChem,
2002, 3, 874
89
Approach B
Generating the Nature Mimics
Direct assay
Cox, R.J., Gibson, J.S., Martín, M.B.M.,
ChemBioChem, 2002, 3, 874 Blanco, J., Moore,
R.A., Viola, R.E., PNAS, 2003, 100, 12613 Han,
S., Moore, R.A., Viola, R.E., Synlett, 2003, 6,
845
90
Biosynthesis of Saturated Fatty Acids
FabI / K / L
ACP
FabD
FabZ
FabH
FabG
Campbell, J.W., Cronan, J.E.Jr., Annu. Rev.
Microbiol., 2001, 55, 305
91
Biosynthesis of Saturated Fatty Acids
FabI / K / L
Continues...
FabD
FabZ
FabF
FabG
Campbell, J.W., Cronan, J.E.Jr., Annu. Rev.
Microbiol., 2001, 55, 305