145

336

Isoprenoid Natural Products: Chapter 4 in Mann, pp 31-52

Terpenes- C10 natural products derived from geranyl pyrophosphate

(2 isoprene units)

Sesquiterpenes- C15 natural products derived from farnesyl pyrophosphate

(3 isoprene units)

Diterpenes- C20 natural products derived from geranylgeranyl pyrophosphate

(4 isoprene units)

C10, C15, C20, etc. natural products derived from “isoprene units”C5 unit Ruzicka isoprene rule

OPP

OPP

OPP

337

Sesterpenes- C25 natural products derived from geranylfarnesyl pyrophosphate

(5 isoprene units)

Steroids & Triterpenes- C30 natural products derived from squalene (2 franesyl units joined head-to-head)

Carotenoids- C40 natural products derived from phytocene(2 geranylgeranyl units joined head-to-head)

OPP

146

338

Coenzyme A (CoASH)

N

N

N

NH2

N

O

OP

O

OH

O P O

OH

OH

O

HN

HN

O

HS

OHO

OH

PHO

O

O

Biosynthesis of Acetyl CoA

N

SPPO

R

OH

CH3

pyruvate decarboxylase

S

NH

O

Lys

S

H+SH

NH

O

LysSN

S

R H3C OH

B:

- thiamin-PP

SH

NH

O

SH3C

O

Lys

CoA-S -SH

NH

O

LysSCoAS

H3C O-

S-CoA

O

SH

NH

O

LysHS+

O

CH3C CO2H

from glucose

thiamin

- CO2

S

NH

O

Lys

S

FAD FADH-

PPO

acetyl CoA

OH

OH

O

HNHO

O

Pantothenic Acid(vitamin B5)

339

Biosynthesis of mevalonic acid:

C

O

H2C SCoA

HB:

H2C SCoAC

O

NHHN

S

O

CH4CO2H

O

COHO

+NHN

S

O

CH4CO2H

HO

O

H+

Biotin Carboxybiotin

SCoA

O

HO2C

malonylCoAacetyl CoA

SCoA

O

malonylCoA

O

OH

B:H2C SCoA

C

O

C

O

H3C SCoA

acetyl CoA

SCoA

OOClaisen

condensation

Acetoacetyl CoA

-CO2

147

340

Biosynthesis of mevalonic acid (con’t):

H

R

O

OO

O

R=H, compactinR=CH3, mevinolin (Lovastatin)

HMG-CoA reductase inhibitorsCholesterol lowering agents

SCoA

O

HO2C

-CO2

H2C SCoAC

O

SCoA

OO

SCoAHO2C

OHH3C O

3-Hydroxy-3-methylglutaric acid(HMG-CoA)

HMG-CoAreductase

NADPH OHHO2C

OHH3C

Mevalonic acid

ATP AMP

O

OHH3C

P O

O-

O

P O-

O

O-

O

O

H

B:

H+

O P O

O-

O

P O-

O

O-

rearrangment

O P O

O-

O

P O-

O

O-HS

HR HS

dimethylallyl-PP(DMAPP)

- CO2

isopentenyl-PP (IPP)

B:

H+

341

OPPOPPHR HS

dimethylallyl-PP(DMAPP)

isopentenyl-PP (IPP)

tail tailhead head

electrophilic head nucleophilic tail

OPP

Biosynthesis of geranylpyrophosphate

148

342

F3C OPP

-CF3 group is predicted to destabilize a carbocation intermediate

Rate was found to be 106 slower

OPP

OPPHR HS

OPP

OPP B:

geranyl pyrophosphate

OPP

OPP

OPP

HR HS

B:

OPP

geranyl pyrophosphate (C10)

farnesyl pyrophosphate (C15)

OPP

geranylgeranyl pyrophosphate (C20)

IPP

etc

343

Monoterpenes: essentials oils (essences)

O

O

Camphour

(+)-Carvone (caraway seeds)(-)-Carvone (spearmint)

(+)-limonene (oranges)(-)-limonene (lemons)

!-pinene "-pinene

CO2H

Chryanthemic acids

OH

Grandisol

149

344

Conversion of geranyl-PP to terpenes involves a cyclization ofan alkene onto a carbocation

OPP

OPP

OPP

Linallyl-PP

OPP

H2O

OH O

Camphour

Can notcyclize

Biosynthesis of camphour

345

OPP

Linallyl-PP

OPP

H2O

OH

!-terpineolcamphene

H

1,2-hydrideshift

H

H

1,2-hydrideshift

H H

O1) H2O2) oxidation

thujone

150

346

Double bonds position of α- and β-pinene

OPP

PPO

!-pinene

"-pinene

OPP

!-pinene

"-pinene

- H+

347

Squalene Biosynthesis: head-to-head coupling of two farnesyl-PPunits catalyzed by squalene synthase

OPP

OPP

S-enzyme

HH

B:

CH3

H

NADP-H

CH3

HOPP

PPO

- OPP+

S-Enzyme_

+

Squalene

151

348

O P O P O -

O

O -

O

O -

O

H A

HO

HO

Farnesyl pyrophosphate (C15)

Squalene Oxide (C30H50O)

2

Squalene synthetase

Squalene (C30H50)

+

Squalene epoxidase

+

Squalenecyclase

Protosterol (C30H50O)

HO

+

HO+ HO +

H

CH3

H3C

HO

CH3

CH3

CH3H

H

H

H

P450

Steroid Biosynthesis: Polyene cyclization

349

H A

HO HO

Cholesterol (C27H46O)Lanosterol (C30H50O)

Protosterol (C30H50O)

CH3

H3C

HO

CH3CH3

CH3H

H

H

H CH3

H3C

HO

CH3CH3

CH3H

H

H

H

+

CH3

H3C

HO

CH3

H

CH3

CH3

H

Lanosterol (C30H50O)

152

350

Biosynthesis of abietic acid

HO2C

head-to-head ?

OPP

H+

OPP - H+

OPP

- H+

CH3 - H+

CH3

H+CH3

1,2-methylshift

abieticacid

351

Stereochemistry of the ring juncture in isoprenoid biosynthesis(polyene cyclizations)

Stork-Eschenmoser Hypothesis: olefin geometry (usually trans) ispreserved in the polyene cyclization reaction. E-olefins lead to trans-fused ring junctions.

OPPOPP

E

E

H

trans-fusedring

O

Protosterol (C30H50O)

CH3

H3C

HO

CH3CH3

CH3H

H

H

H

squalene epoxide

E E

E

trans-fused ring

153

352

Stork-Eschenmoser Hypothesis: conformational analysis of the polyene cyclization reaction

Transition state leading to the cis-fused product is disfavored

H3C

H

R

CH3

H

CH3

CH3

+

CH3

CH3

CH3H3C

R

HH

trans-fused

H3C

CH3

CH3

+H

H

HH3C

RH

H

1,3-diaxial interactions

H3CCH3

CH3

+H

H

HH3C

R H

H

H

cis-fused

353

in vitro polyene cyclization

O O

SnCl4pentane

H

HH

H

OHO

(27%)

W.S. JohnsonJACS 1974, 96, 3979

O

H

H

H H

HO

SnCl4CH3NO2, 0°C

!- Amyrin

E.E. van TamelenJACS 1972, 94 , 8229

(8%)

154

354

OO O O

O

OH

HO

H3CO

HO2

C

OH

OO

H OH

HOOH

HO

HO2C

O

O

O

O

O

O

OOH

H

H

M+

monensin A

lasalocid A

O

O

OH

O

O

O O

O

HON(CH3)2

CH3

OH

OCH3

CH3

O

H3CO

O

O

O

CHO

O

O

O OOHO

N(CH3)2

O O O

HO

erythromycinCarbomycin A

OCH3 O

O OH

OH

O

OH

O

H3C

OHNH2

daunomycin

CH3

OH

CO2H

6-methylsalicylic acid

Polyketide Derived Natural Products:

355

N

N

N

NH2

N

O

OP

O

HO

O P

O

OH

OH

O

HN

HN

O

HS

OHO

HO

P

HO

O

O

CoA

serine-proteinO P

O

OH

OH

O

HN

HN

O

HS O

4'-phosphopantethienyl group

Polyketide Biosynthesis: acyl carrier protein

155

356

O

CH3C SCoA ACPHS+

O

CH3C S ACP+ CoA-SH

O

C-O2CH2C SCoA

acetyl-CoA

malonyl-CoA

ACPHS+

O

C-O2CH2C S ACP

+ CoA-SH

O

CH3C S ACP +O

C-O2CH2C S ACP

O

CCH2 S ACPC

O

H3C

Claisencondensation

elongation

CH3

CO2H

OH

aromatics(phenols)

CO2H

full reduction, then elongation

fatty acids

O

O

OH

O

O

O O

O

HON(CH3)2

CH3

OH

OCH3

CH3

partial reduction,than elongation

acetate/propionate(macrolides)

357

Chain Elongation: Fatty acid biosynthesis Chapter 2 of Mann, pp 10-18

ACP

SH

ACP

SH

Cys

SH

Cys

SH

ACP

S

Cys

S

O O

O -

O

-CO2

ACP

S

Cys

S

O O

ACP

S

Cys

SH

O O

NADPH

ACP

S

Cys

SH

O OH

HR HS- H2O

ACP

S

Cys

SH

O

HR NADPH

ACP

S

Cys

SH

O

ACP

S

Cys

S

O

O -

O O

!-ketoreductase dehydratase

enoyl reductase

!-ketosynthase

156

358

Aromatic (phenolic) polyketide biosynthesis:

ACP

S

Cys

S

O O

O -

O

-CO2

ACP

S

Cys

S

O O

ACP

S

Cys

SH

O O O

O

ACP

S

Cys

SH

O O O

ACP

S

Cys

SH

O O O O

etc.

359

Cylization and aromatization of the polyketide:6-methyl-salicylic acid

O

O

S ACP

ONADPH

HO

O

S ACP

O- H2O

O

S ACP

O

O2C COSCoA

O

O

S

O

ACP

O

O

S

O

ACP

OH

O O

S ACP

- H2O

O O

S ACP

tautomerization

OH O

S ACP

H2O

OH O

OH

157

360

O

OH

OH3CO

O

O

OH

OH

O

HO

H3C

NH2

daunomycin

SAM

hydroxylation/oxidation

O

O O O O

O O O

SO ACP

O

proprionate

lost as CO2

oxidation

Not the same as on page 30 in Mann

361

OCH3

O

O

OH

OH

O

O

OH

OH3C

OHNH2

Daunomycin

O

H3COH

H3C

OH

O

CH3

H3C

OR2

OR1

CH3

O

CH3

OH

Erythromycin

OO O

CH3

H3CO

CH3

NaO2C

H3C

HO

CH3

OO

H3C H3C

CH3

HOOH

H3C

HO

CO2Na CH3

OH

CH3

OO

O

CH3

OH

H3C

Monensin A

Lasalocid A

acetate propionate butyrate

Polyketides:

158

362

Mapping the carbon skeleton by using 13C-labeled precursors

O

H3C OH

H3C

OH

O

CH3

H3C

OR2

OR1

CH3

O

CH3

OH

Erythromycin

O -

O

•• = 13C

••

•• •

••

O

H3C OH

H3C

OH

O

CH3

H3C

OR2

OR1

CH3

O

CH3

OH

O -

O

O -

O

•

••

••••

•

O

OH

OH

O OR2

OR1

O

OH••

••••

•

••O SCoA

•

•

• •

••

•

363

Origin of the oxygen atom in macrolide biosynthesis

CO2H

CH3H

HS1O

CH3H

HS2O

HH

HH3C

O

CH3H

OHH3C

OHH3C

OHH

CH2CH3

HHO

1

9

O

OH

OH

O OR2

OR1

O

OH

1

9

159

364

Three hypotheses for the origin of the oxygen atoms:

SCoA

O

O

OH

OH

O OR2

OR1

O

OH

COSCoA COSCoA

OH

OH

OH

OH

O

O2

SCoA

OH

OH

O

O

OH

OH

O OR2

OR1

O

OH

SCoA

OH

OH

O

SCoA

OH

O

SCoA

OH

OH

O

H2O

365

18O Isotope Shift of the 13C Resonances

C18O H

H3C

H3C

H3CO

18O

HO

HOHO

HO

H

160

366

O

H3COH

H3C

OH

O

CH3

H3C

OR2

OR1

CH3

O

CH3

OH

Erythromycin

O -

O

18O

The other two oxygenation come from O2, so they presumablycome from a P450 mediated hydroxylations.

Origin of the oxygen atoms:

367

Origin of the Oxygen Atoms in Monensin Biosynthesis

H3C

O

O-

H3CH2C

O

O -

H3C

O

O-

H3CH2C

O

O-

18O2

OO O

CH3

H3CO

CH3

NaO2C

H3C

O

H3C H3C

CH3

HO OH

HO

CH3

O

O O O

CH3

H3CO

CH3

NaO2C

H3C

HO

CH3

OO

H3C H3C

CH3

HO OH

S. Cinnamonensis

18O labeled

acetate and

propionate

S. Cinnamonensis

161

368

Origin of the Oxygen Atoms in Monensin Biosynthesis

[O]

OH

O

CH3

H3CO

CH3

NaO2C

H3C

HO

O

OH

O

CH3

H3CO

CH3

NaO2C

H3C

HO

O

O

O

O

H+

acetatepropionatebutyrate

Monensin

369

Biomimetic synthesis of polyethers based on polyepoxide cyclizations

O

H3COO O

O

H

PLE

50 mM phosphatepH 8.0

O O O OHOCH3 CH3

O

O

O O

O

AcOH

O O OO

CH3 CH3

OH

OH

162

370

Polyketide synthase (PKS) modules:

Khosla, C. Chem. Rev. 1997, 97, 2577-2590.

6-Deoxyerythronolide B synthase (DEBS)

371

Genetic manipulation of PKS modules as a means of generating newnatural products:

163

372

OH

OH

O

O

O

CHO

tylosin aglycone

H3CO

OH

OH

OH

O

O

CHO

spiramycin aglycone

HO

373

Alkaloid Biosynthesis

Alkaloids: non-amino acid, non-nucleoside, non-cofactor, natural product that contains a basic nitrogen

Where does the nitrogen come from?

Alkaloids

Amino acid Polyketide or Isoprenoid

+ NH3

164

374

Some Alkaloids

N

N

CH3

nicotine

NH2

OCH3

H3CO

H3CO

mescaline

N

H3C

CO2CH3

O Ph

O

cocaine

N

O

N

O

H

H

H

H

strychnine

O

HO

N

CH3HH

HO

morphine

N

NH

HO2C

H

CH3

lysergic acid

NH

N

HO

H3CO

H

quinine

NH

N

HO

MeO2C

NMeO

N

H

OH

CO2CH3

O

O

R

R= -CH3 vinblastine

R= -CHO vincristine

N OH

H3C

Lycopodine

NH

OH

Histrionicotoxin

375

Amino Acid Precursors

N

N

CH3

nicotine

from

ornithine NH2

OCH3

H3CO

H3CO

mescaline

fromtyrosine

N

H3C

CO2CH3

O Ph

O

cocaine

from

ornithine

N

O

N

O

strychnine

from tryptophan

O

HO

N

CH3

HO

morphine

fromtyrosine N

NH

HO2C CH3

lysergic acid

from tryptophan NH

N

HO

H3CO

H

from tryptophan

NH

N

HO

MeO2C

NMeO

N

OH

CO2CH3

O

O

R

R= -CH3 vinblastineR= -CHO vincristine

from tryptophan

N OH

H3C

Lycopodine

from lysine

NH

OH

Histrionicotoxin

165

376

Note: amino acid precursors are lysine, ornithine (arginine),tryptophan, tyrosine

pyrrolidine rings are derived from ornithine, not proline

Alkaloids derived from lysine and ornithine (arginine)

HO2C

HN NH2

NH2

NH2

arginine

H2O

HO2CNH2

NH2pyridoxal

phosphate

H2NNH2

ornithine putrescine

HO2C NH2

NH2

pyridoxalphosphate

H2N NH2

lysine cadaverine

- CO2

- CO2

377

Alkaloids derived from ornithine: Biosynthesis of Cocaine:

H2NNH2

putrescine

NH

NH2SAM H3C

pyridoxalphosphate

NH

OH3C

H

-H2O

N

CH3

O2C

O

SCoA

N

CH3

O

SCoA- CO2

O2C

O

SCoA

- CO2

N

CH3

O

SCoA

O P450

N

CH3

O

SCoA

O

HO

-H2O

N

CH3

O

SCoA

OH

N

H3C

O

SCoA

O

166

378

Biosynthesis of Cocaine

N

H3C

O

SCoA

O

N

H3C

O

OH

O

-H2O

SAM

N

H3C

O

OCH3

O

N

H3C

OH

OCH3

O

NADPH

N

H3C

O

OCH3

O

Ph

OCocaine

379

Alkaloids derived from ornithine: Biosynthesis of Nicotine

NH

CO2HNADPH

NH

H

H

O

O

NH

H

H

N

CH3

NH

N

CH3

H NADPH

from ornithine

NH

H

H

N

CH3

NADP+

Nicotinic Acid

+

- CO2

H

167

380

Alkaloids derived from lysine: piperidine containing alkaloidsin analogy to the ornithine derived alkaloids

H2N NH2

cadaverine

NH

O N

H3C

CH3

N

CH3

Nu

Nu:

NPh

OH

Ph

O

CH3

(-)-lobeline

NH

O

pelletierine

NH

coniine

NH n

Fire ant venomn= 10, 12, 14

Not derived from lysine, but rather from polyketide biosynthesis

381

Proposed biosynthesis of coniine:

O

NH

coniine

CoAS O

O

O

CoAS

O

CO2

CoAS

O+

CoAS O

OH O

O

NADPH

alanine

transaminase

NH2

O

NH

- H2O

NADPH

168

382

N

HO OH

retronecine

derived from two ornithine units

pyrrolizidine alkaolid quinolizidine alkaolid

N

OH

Lupinine

derived from two lysine units

N

N

Sparteine

derived from three lysine units

383

Some details of pyrrolizidine alkaloid biosynthesis

H2NNH2

putrescine

2 H2NNH2

ONH2+

H2NN

NH2

- H2O

H2N

HN

NH2

NADPH

homospermidine

N

HO HOH

N

NH2

PAL

PAL

O

HN

NH2

- H2O

N

H CHO

N

HOH

retronecine

169

384

Alkaloids derived from tyrosine. Morphine Biosynthesis

NH2

HO

H

OHO

-CO2

-CO2

CO2H

NH2HO

CO2H

OHO

PAL

PAL

thiamin

hydroxylation

Tyramine

NHHO

HO

HO

NH2

HO

HO

Dopamine

+NH

HO

HO

HO

2 SAMN

HO

H3CO

HO

CH3

Norcoclaurine

385

NHO

H3CO

HO

CH3

1) hydroxylation2) SAM

NHO

H3CO

H3CO

CH3

HO

Reticuline

NHO

H3CO

H3CO

CH3

HO

HO

H3CO

N

CH3

OH

H3CO

epimerization

"- 2 H•"

•O

H3CO

N

CH3

O •

H3CO

O

H3CO

N

CH3

O

H3CO

HO

H3CO

N

CH3

O

H3CO

••

170

386

HO

H3CO

N

CH3

O

H3CO

NADPHHO

H3CO

N

CH3

OH

H3CO

O

H3CO

N

CH3

H3CO

1) P4502) isomerization3) NADPH

O

H3CO

N

CH3

HO

O

HO

N

CH3

HO

CodeineMorphine

P450

387

Alkaloids derived from tryptophan. Physostigmine biosynthesis:

NH

NH2

CO2H PAL

NH

NH2

tryptophan tryptamine

SH3C

asenosyl

R

NH

NH2

CH3

NH

CH3

NH

N

CH3

N

CH3 CH3

OO

NHH3C

physostigmine

171

388

Alkaloids derived from tryptophan. Lysergic acid biosynthesis:

NH

NH2

CO2HOPP

NH

NH2

CO2H

SAM

NH

HN

CO2H

CH3

[O]

NH

HN

CO2H

CH3

OH

-H2O

NH

HN

CO2H

CH3

NH

O

HN CO2H

CH3

NH

HNCH3

HO

-CO2

NH

HN

CHO CH3

PAL1) [O]2) rearrangment

P450

389

Lysergic acid biosynthesis (con’t):

NH

HN

CHO CH3

- H2O

NH

NCH3

NH

NCH3HO2C

1) [O]2) rearrangment

Lysergic Acid

NADPH

NH

NCH3