Polysaccharides Glycosides Nucleic Acids Shikimate pathway Aromatic Compounds Lignans aromatic amino...

PolysaccharidesGlycosidesNucleic Acids

Shikimate pathwayAromatic CompoundsLignans

aromatic amino acidsaliphatic amino acids

CH3COSCoA

CH3COSCoA

(photosynthesis)

phosphoenol pyruvate

pyruvate

acetylCoA CH3COSCoA

-O2CCH2COSCoA

CH3COSCoA

Polyketides, Fatty Acids Prostaglandins, Macrocyclic Antibiotics

Shikimate

CH3COCH2COSCoA

mevalonate

Isoprenoids (terpenes,steroids, carotenoids)

AlkaloidsPeptidesPenicillinsCyclic Peptides

CITRICACIDCYCLE

(3)

(3) Mevalonate Terpenes, Steroids

-- made from 5-carbon units

-pinene (C10) cholesterol (missing 3 C’s)

phytane (C20)

(3) Isoprene pathway: Terpenes

- C6 compound that loses CO2

to form C5 units

Isoprene Biosynthesis

DMAP IPP

- Result: two isomeric 5-carbon molecules, IPP + DMAP

- Pyrophosphates: high-energy group powers biosynthetic rxns.. “nature’s leaving group”

decarboxylates to yield IPP

Biosynthesis of Monoterpenes (C10)

rearranged to form sesquiterpenes (C15)

Geranyl pyrophosphate (C10)

Farnesyl pyrophosphate (C15)

Prenyl transferase +

DMAP IPP

-OR-

Cyclase enzymes

cyclic monoterpenes (C10)

Biosynthesis of Monoterpenes (C10)

(1) DMAP ionizes to form electrophilic carbocation

(2) Nucleophilic attack by IPP forms geranyl-PP

(3) Stereospecific loss of HR, forming double bond

(4) Geranyl-PP ionizes, rearranges to form a carbocation intermediate

- Cyclic monoterpenes then form via enzyme-catalyzed stereospecific rearrangements, functionalizations

thujone

(+)-carvone = caraway seed(-)-carvone = spearmint

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

Most cyclic monoterpenes have a distinctive odor - basis of perfume & flavor industries

Stereoisomers have different characteristic smells - demonstrates that smell receptors are 3D proteins, i.e. chiral environments that can distinguish enantiomers

Biosynthesis of Sesquiterpenes (C15)





DMAP IPP

Biosynthesis of Sesquiterpenes (C15)





DMAP IPP

- if you introduce a labeled carbon in the precursor, you can see where it ends up in the final natural product

Labeling studies in Biosynthesis 1

Using isotopically enriched precursors, it is possible to trace the incorporation of building blocks into complex metabolites

- follow the label and see which atoms in the natural product end up labeled

Originally done with radioactive labels (14C, 3H)

- feed cells a precursor with one 14C

- use chemical degradation to break natural product apart, see which pieces contain the radioactivity

- drawback: sloppy, usually not possible to completely determine biosynthetic route

Labeling studies in Biosynthesis 2Now done by NMR using non-radioactive isotope 13C

- common carbon isotope 12C is invisible to NMR

- rare 13C has nuclear spin I = n/2, so is detectable by NMR

- feed cells a precursor with one position 13C labeled; then see which positions show exaggerated peaks in the carbon NMR spectrum

since only 1% of carbons are naturally 13C, the positions that incorporate the label will give much higher peaks

13C NMR spectrum of compound 2, after feeding sea slug with [2-13C] mevalonate - increased size of peaks shows label incorporation

Kubanek et al. JOC 1997

- allowed researchers to infer biosynthetic pathway for these related C-23 and C-21 terpenes

Trichodiene

- Precursor (mevalonate) can be labeled at a particular atom with a stable isotope (13C); fate of label then traced in final product

Biosynthesis of Trichodiene (C15)

Trichodiene

- Biosynthesis proceeds via carbocation intermediates, migration of 2 methyl groups and a hydrogen

Biosynthesis of Trichodiene (C15)

Trichothecenes

Trichodiene is a key precursor of fungal metabolites called trichothecenes, a major human health problem

- fungal toxins are called mycotoxins - often present in moldy grain consumed by cattle or people

Winter 1942: thousands of Russians died after eating grains left in the field over winter, due to the war; grain was infected with the trichothecene-producing fungus

1981: USA accused Russia of spraying fungal spores in Vietnam, resulting in foliage contaminated with trichothecenes (false)

Trichodiene

Diterpene (C20) Biosynthesis


+

Gerenylgerenyl-PP (C20) carbocationintermediate

Diterpenes

Gibberellin Biosynthesis

CH3 oxidized, then lost as CO2

Gibberellin A3, a potent C19 plant hormone

- found in most plants (though originally isolated from a fungus)

DMAP IPP

Squalene (from shark oil)

C30 compound: 2 farnesyl’s joined tail-to-tail



Triterpene (C30) Biosynthesis

Triterpene (C30) Biosynthesis

Squalene

other C30 triterpenes

cholesterol (C27)

sex hormones

vitamin D

Cholesterol Biosynthesis

- lose 3 methyl groups

Vitamin D Biosynthesis

- cholesterol from liver is transported to skin

- photochemically converted into vitamin D- vitamin allows uptake

of essential calcium

Notes on Terpenes(1) Oxidation reactions are carried out by the enzyme cytochrome P450

- activate oxygen to introduce -OH, carboxyl groups- allow removal of C’s through decarboxylation

(2) Triterpenes form flexible rings (chair, boat conformations) with many chiral centers {rings usually not aromatic}

- provides a huge number of potential 3D structures- high degree of biological activity

(3) Pathways can be elucidated using labeled precursors, such as mevalonate with a 13C at position 2

- carbon NMR experiments reveal where the label ends up in the completed molecule

Higher terpenes (C40)

Lycopene

2 x geranyl geranyl-PP

- major antioxidant pigment in tomatoes

-carotene- major accessory pigment in photosynthesis

The highest terpenes (Cbig #)~ 1 % of plants can synthesize cis-polyisoprenoids, like rubber

H3C

Commercially used rubber plants can convert nearly 100% of their mevalonate into rubber

PolysaccharidesGlycosidesNucleic Acids

Shikimate pathwayAromatic CompoundsLignans

aromatic amino acidsaliphatic amino acids

CH3COSCoA

CH3COSCoA

(photosynthesis)

phosphoenol pyruvate

pyruvate

acetylCoA CH3COSCoA

-O2CCH2COSCoA

CH3COSCoA

Polyketides, Fatty Acids Prostaglandins, Macrocyclic Antibiotics

Shikimate

CH3COCH2COSCoA

mevalonate

Isoprenoids (terpenes,steroids, carotenoids)

AlkaloidsPeptidesPenicillinsCyclic Peptides

CITRICACIDCYCLE

(4)

(4) Acetate Polyketides

H3C SCoA

O

Erythromycin A (antibacterial)

Avermectin B1

(antihelminthic)

(4) Polyketide Biosynthesis

(4) Acetate Polyketides:

(1) aromatic compounds, usually with meta placement of -OH’s

(2) non-aromatic macrolides H3C SCoA

O

Avermectin B1

(antihelminthic)

(4) Polyketide Biosynthesis

6-methyl salicylate

Polyketide BiosynthesisCompounds constructed by addition of successive 2-carbon units

Ketone may or may not be later reduced

Options in final structure:

H3C C

O

H3C C

OH

HH3C C

-or- -or-

Polyketide BiosynthesisImportant natural producers of polyketide metabolites:

- fungi (= molds)

- bacteria, particularly of the family Actinomycetes

- sea slugs

Polyketide BiosynthesisActual pathway starts with an acetyl-coA, but then successively uses malonyl coA, which loses CO2 thus adding C2 units

- analogous to how mevalonate loses CO2 to add C5 units in terpenoid biosynthesis

1 acetate, 3 malonyl coA’s

4 carbons are labeled in the final product

The Lactone Mellein[1,2-13C]-acetate

From the fungus Aspergillus

Growing polyketide chains are held bound to the biosynthetic enzyme, passed from one active site to the next

Different active sites carry out the various cyclizations + reductions

The final site is a thioesterase, which cleaves the connection, setting the compound loose

Griseofulvin Biosynthesis

Start with 14 carbons in linear precursor…

…End with 17 ?

Griseofulvin Biosynthesis

Post-cyclization modifications: (1) addition of methoxy groups (2) halogenation

Actinorhodin Biosynthesis

actinorhodin

Antibiotic from bacterium Streptomyces coelicolor

kalafungin(an antibiotic intermediate)

methylated afterwards

Aflatoxin

Isolated in 1960 after thousands of turkeys died of liver failure from eating moldy peanut meal; culprit was Aspergillus mold

Now evidence that in parts of the world where peanuts are a dietary staple, much liver disease is due to persistent aflatoxin consumption

Erythromycin

Synthesized using propionate, C3 units, instead of 2-carbon acetate units (hence all those methyl groups)

Antibiotic from bacterium Streptomyces erythreus

6-deoxy-erythononlide B

Erythromycin

2 sugars added last, not part of the polypropionate pathway

cladinose

desosamine

Erythromycin

Biosynthetic genes for erythromycin have been cloned: - made by 3 large, polyfunctional proteins called modular polyketide synthases

- proteins have a series of similar active sites that carry out each successive step in the biosynthesis (hence “modular”)

Mixed Biosynthesis: Vitamin K

Synthesis performed by enteric bacteria in large intestinePart of vitamin is from shikimate pathway, part isoprene

isoprene unit

Rules for identifying pathways(1) Is there nitrogen? Yes... alkaloid

(2) Count carbons a) multiple of 5.... terpene

C10 = monoterpene C15 = sesquiterpene C20 = diterpene

b) another even number... probably polyketide

c) multiple of 3... polypropionate (w/ lots of methyls)

especially if the rings are not aromatic

Rules for identifying pathwaysHints for carbon-counting:

1) non-skeleton carbons don’t count - Those attached through oxygen (esters, ethers) are usually (but not always) added after the main pathway

- For example, in R-OMe, the methoxy carbon would not be included in your count (secondary modification)

2) sugars do not count; they are often tacked onto shikimate, polyketide metabolites after the synthesis is complete

Rules for identifying pathways(3) Aromatic...

a) hydroxyl groups are meta... polyketide

b) otherwise... shikimate

You can’t always differentiate these 2 pathways without doing labeling experiments

Polysaccharides Glycosides Nucleic Acids Shikimate pathway Aromatic Compounds Lignans aromatic amino...

Documents

Transcript of Polysaccharides Glycosides Nucleic Acids Shikimate pathway Aromatic Compounds Lignans aromatic amino...