Traditional approach for bioactive natural product discovery

fractionation

extraction

Investigate bioactivity of extract

Identify active fraction(s) and determine structure(s) of compound(s)

Genome sequence of the model antibiotic-producer Streptomyces coelicolor M145

NH

N

HN

OMe

prodiginines

OO

O

O

OH

OH

OH

OHO

O CO2H

HO2C

actinorhodin

NH O

O HN

O

HN

HN

CO2H

O NH

O

HN

ONH

OOH

NH

OHN

R'HO2C H

NO

OH2NOC

NH

O

HN

O

NH

O

CO2H

OH

O

HO2C

OR

calcium-dependent antibiotic

O

CO2HO

methylenomycin A

Nature (2002) 417, 141-147

Cryptic biosynthetic pathways encoded within the S. coelicolor genome

DEBS – an archetypal cis-AT polyketide synthase

ACPAT

Load Module 1

DEBS1

SO

ACPKS

KR

SO

HO

AT TEACPKS

KR

SO

HO

AT

HO

Module 2 Module 3

DEBS2

ACPKS

SO

O

AT ACPKS

SO

AT

O

Module 4

KRDHER

Module 5

DEBS3

ACPKS

KR

SO

HO

AT ACPKS

KR

SO

HO

AT

HO

Module 6

HO

HO

O

HO

HO

O

HO

HO

HO

HO

OH

O

O

OH

OHO

6-deoxyerythronolide B

HO

HO

O

HO

HO

O O

O

O

OHOH

O

OH

O

O

O

O OH

OMe NMe2HO

erythromycin A

O

SCoA

O

SCoA

O

-O

+

Nature (1990) 348, 176; Science (1991) 252, 671

ACPKS

DH

ER

KR

AT

OH

ACPKS

DH

ER

KR

O

SH

AT

Posttranslational ACP modification

CoASH

ACPKS

DH

ER

KR

SCoA

O

O-

O

AT

OH S-

Acyl transferase

KS

DH

ER

KR

OH S

O

O-

O

AT ACPACPKS

DH

ER

KR

O

O-

O

AT

O S-

Prediction of AT domain substrate specificity

Haydock et al. FEBS Lett. (1995) 374, 246-248Banskota et al. J. Antibiot. (2006) 59, 168-176

Malonyl-CoA

Methylmalonyl-CoA

ACPKS

DH

ER

KR

S- S

O

O-

O

AT ACPKS

DH

ER

KR

S S

O

O-

O

O

R

AT

Ketosynthase 1

ACP

S

O

R

ACP

S-

Ketosynthase 2

ACPKS

DH

ER

KR

S S

O

O-

O

O

R

AT ACPKS

DH

ER

KR

S- S

O

R

O

AT-CO2

Keto reductase

ACPKS

DH

ER

KR

S

O

R

HO

ATACPKS

DH

ER

KR

S

O

R

O

ATNADPH

Prediction of KR domain stereospecificity

Keatinge-Clay, Chem. Biol. (2007) 14, 898-908

Dehydratase

ACPKS

DH

ER

KR

S

O

R

ATACPKS

DH

ER

KR

S

O

R

HO

AT-H2O

Enoyl reductase

ACPKS

DH

ER

KR

S

O

R

ATACPKS

DH

ER

KR

S

O

R

ATNADPH

Prediction of ER domain stereospecificity

Kwan et al., Chem. Biol. (2008) 15, 1231-1240

S

O

S

O-

S

ONADPH H+H

H

H

S

O

H

HorS R

Thioesterase 1

ACP

OH

TE

S

O HO

ACP

SH

TE

O

O O

H

ACP

O

TE

S-O HO

Thioesterase 2

ACP

SH

TE

O-O O

ACP

SH

TE

OH

O

O

ACP

SH

TE

O

O O

H