Natural Products 2

7/28/2019 Natural Products 2

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UPTAKE OF NITROGENBY PLANTS


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THE NITROGEN CYCLE

NitrogenFixingBacteria

N2

AminoAcids

HigherPlants

HigherPlants

NO3- NH3

NO2-

NitrifyingBacteria(Nitrobacter)

NitrifyingBacteria(Nitrosomas)

proteins, nucleic acidselectron carriers,alkaloids

returned tothe soil or the

atmosphere

ETC


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In this section we will study the mechanismsby which plants take up nitrogen.

First, however, we need to look at a couple ofcoenzyme systems.

NAD+ NADH

NADP+ NADPH


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THE NAD+ / NADP+ COENZYMES


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N

NN

NN

CNH

2

O

O

CH2

O P O

O

O

P

O

O

O CH2

O

OH OH

OH O

NH2

R- -

+

NICOTINE ADENINE DINUCLEOTIDE

NAD+

NADP+

R = -H

R = -PO3H

N

CNH

2

O

R+

Natures HydrideReducing / OxidizingCoenzyme (reagent)

membranes

cytosol

The two forms differby a phosphate groupwhich also controls the

location in the cell.


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NAD+ and NADP+ ARE HYDRIDE ACCEPTORS

OXIDATION

N

C

NH2

O

R+

C

O

H

H

C

O

N

C

NH2

O

R

H

:

H

..

hydride transfers

REDUCTION

:B-EnzH-B-Enz

NAD+

NADP+

NADH

NADPH

NADH and NADPH ARE HYDRIDE DONORS

alcohol ketone oraldehyde

Unlike ordinary chemical reagents,these coenzymes function reversibly.

AROMATICBUT

CHARGED

UNCHARGEDBUT NOT

AROMATIC

OXIDATION

REDUCTION

-2e-

+2e-

-2e-

+2e-


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N

CNH

2

O

R+

N

CNH

2

O

R

H

H

..

hydride

acceptor

NAD+

NADP+

NADH

NADPH

Unlike ordinary chemical reagents,these coenzymes function reversibly.AROMATIC

BUTCHARGED

UNCHARGEDBUT NOT

AROMATIC

OXIDATION

REDUCTION

-2e-

+2e-

H:-

BIOCHEMICAL REVERSIBILITY

hydride

donor

Many biochemical reagents aredesigned to reversible -

this is the ultimate in conservation


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C

O

H

H

C

O

NAD+ NADH

BIOCHEMICAL NOTATION

C

O

H

H

C

O

NADH NAD+

oxidation

reduction


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OTHER FUNCTIONAL GROUPS CAN BE OXIDIZED / REDUCED

C

N

H

H

H

C

NH

NADH NAD+

reduction

imine amine

:..

C

NH

N

CNH

2

O

R

HH

..

Enz-B-H

:

This process is one ofthe steps of

REDUCTIVE AMINATION

one of the ways thatplants take upnitrogen.


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REDUCTIVE AMINATION

C O

H3

N:

H-B-Enz

..: C O HNH

H- H2O

N CH

N

CNH

2

O

R

HH

..

NAD+

Enz-B-H

NADH

CNH

H

..

....

aldehydeor ketone

imine

amine


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C

O

RR R C R

NH2

H

NH4+ HCO2

-

DC

N

RR

HH

H C

O

O

+

-....

:

..:

formateion

CLASSICAL REACTION

LEUCKART REACTIONREDUCTIVE AMINATION

ammoniumformate


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HOW PLANTS INCORPORATENITROGEN


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GLUCOSE

GLYCOLY

SIS

.a-ketoglutaric acid

CITRICACID

CYCLE

FORMATION OF AMINO ACIDS

CH2

CH2

C

O

COOHHOOC

NH3

NADH

NAD+

CH2

CH2

C

NH2

COOHHOOC

H

an intermediate from

the citric acid cycle

a-ketoglutaric acid

glutamic acid

(NO3-)PRIMARY

METABOLISM

Glutamic acid is the only amino acidthat can be formed by direct uptakeof nitrogen from the soil.

Other amino acids are formed by

transamination.

REDUCTIVE AMINATION


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CH C

OH

R1

O

NH2

CH C

OH

R2

O

NH2

C C

OH

R1

O O

C C

OH

R2

O O

+ +PLP

glutamic acid or

other a-amino acid

NITROGEN EXCHANGETRANSAMINATION

transaminaseenzyme

Most other amino acids are formed by using the -NH2 groupsfrom glutamic acid in transamination reactions.

However, any amino acid can subsequently transfer its amino

group to an a-ketoacid to form a new amino acid.

a-ketoacid

Plants have a severe need for nitrogen and they practice asmuch conservation as possible. Nitrogen is transferred towhere it is needed most.


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CH2

CH2

C COOHHOOC

O

CH2

C

O

COOH

CH3

C

O

COOH

phenylpyruvic acid

pyruvic acid

a-ketoglutaric acid

phenylalanine

CH3

C

NH2

COOH

H

alanine

CH2

C

NH2

COOH

H

CH2

CH2

C

NH2

COOHHOOC

H

glutamic acid

glutamicacid

a-ketoglutaricacid

phenyl-alanine

phenyl-pyruvic acid

alanine pyruvicacid

SOME TRANSAMINATIONS


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FOR MANY OF THE a-AMINO ACIDS FOUND IN NATURETHERE IS A CORRESPONDING a-KETOACID*

alanine pyruvic acid

phenylalanine phenylpyruvic acid

glutamic acid a-ketoglutaric acid

* Some amino acids are made by dimerizations or other non-direct pathways.

valine

leucinea-ketovaleric acid

tyrosine p-hydroxyphenylpyruvic acid

a-ketohexanoic acid

a-amino acid a-keto acid


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CARBAMOYL PHOSPHATE

A Second Method of Transamination

In some plants the major method of nitrogen uptake utilizescarbamoyl phosphate.

NH2

C

O

O P

O

O

OHCO2 + NH4+ + 2 ATP + H2O + 2 ADP + Pi

carbamoyl phosphate

NH2

C

O

O P

O

O

OH

C

O

COOH

.. :Nu C

O

COOH

NH2

H

+ O=C=O

+ Nu-PO3H


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FAD - ANOTHER COENZYME

A Return to Modification of Amino Acids

Process 4


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AMINO ACID METABOLISMWHAT A PLANT CAN DO TO AMINO ACIDS

R CH2

CH

NH2

COOH

R CH2

C

O

HR CH2 C

O

COOH

R CH2

CH2

NH2

C CH

H

R

COOH

1 2

3 4

oxidativedeamination

decarboxylation

oxidative

deamination

decarboxylation deamination

[O]-NH3

[O]-NH3-CO2

-CO2-NH3

Processes 1 2 3 use PLP, pyridoxyl-5-phosphate (B6) coenzymeProcess 4 uses FAD, flavine adenine dinucleotide (B2) coenzyme

and the coenzyme NADH (both discussed later)


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FLAVINE ADENINE DINUCLEOTIDE

N

NN

N

NH2

NH

N

N

NO

O

CH2

CH3

CH3

CH CH CH CH2

O

OHOHOH

P OOH

O

P OOH

O

CH2 O

OH OH

Flavine adenine dinucleotideFAD ( Vitamin B2 )

Cofactor = Mg+2

4

NH

N

N

NO

O


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NH

N

N

NO

O

NH

N

N

NO

O H

H

2e- + 2 H+ +

FAD FADH2

THIS TOO IS A REVERSIBLE COENZYME

It is usually associated with the family of enzymes calledamine oxidases.

R CH2

CH

NH2

COOH

R CH2

C

NH

COOH

R CH2

C

O

COOH

FAD FADH2

amineoxidase

H2O

hydrolysis

imine

a-ketoacid


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POSSIBLE CHEMICAL SEQUENCE FOR DEAMINASES

R CH2

CH

NH2

COOH

R CH2

C

NH

COOH

R CH2

C

O

COOH

FAD FADH2

amineoxidase

H2O

hydrolysis

imine

a-ketoacid

NADH

NAD+

R C C

COOH

O

H

H

H

H

C C

COOH

R H

H

- H2O

dehydration

a,b-unsaturatedacid

a-hydroxyacid

conversion of anamino acid to ana,b-unsaturated

acid

4


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THE MECHANISM FOR FAD OXIDATIONS ANDREDUCTIONS IS NOT COMPLETELY KNOWN

1. It may be a radical-like process involving one-electron transfers.

2. A hypothetical ionic mechanism (mine, not proven) can also be formulated:

NH

N

N

NO

O

C

R

NH2

COOH

H..

NH

N

N

NO

OH

N H

R COOH

H

H

..

Enz-BH

NH

N

N

NO

HOH

..CR COOH

N

HH

+

Enz-B:

CR COOH

NH

NH

N

N

NO

O H

H

+

..

....

Enz-BH

..

..

..


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AMINE OXIDASES

Monoamine Oxidases (MAO)

Diamine Oxidases


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Two types of amine oxidases are common:

Monoamine oxidases and Diamine Oxidases

NH2

COOH

C O

COOH

NH2

NH2

COOH

NH2

O

HCOOH

Monoamine

Diamine

AMINE OXIDASES

.. well see more of this later


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BIOLOGICAL COENZYMES

A Table of Coenzyme Function for Future Reference

The coenzymes are natures chemical reagents and eachhas tis own specific reactions that it can carry out.

Amazingly, nature uses very few coenzymes to carry outthe majority of its chemical transformations.


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COENZYMES - 1

Pyridoxyl-5-phosphate MODIFIES AMINO ACIDS

1) Oxidative deamination to a-ketoacids2) Transaminations3) Oxidative deamination-decarboxylation

to aldehydes4) Decarboxylation to amines

Flavine adenine dinucleotide 1) oxidation of amines to imines(amine oxidases)

2) oxidation of hydrocarbons( alkanes to alkenes )

S-Adenosylmethionine1) Biological methylation reagent for

alcohols, phenols, amines

Nicotine Adenine Dinucleotide

1) Reduction of C=O2) Oxidation of -CH-OH

PLP (P5P)

FAD / FADH2

SAM (SAdM)

NAD / NADP

NADH / NADPH


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COENZYMES - 2

Adenosine triphosphate

ATP

1) Energy storage2) Modifies alcohols3) Traps alcohols4) Coupled reactions

Coenzyme A

CoA

1) Transfers acyl groups

Biotin 1) Carries / transfers CO2 in theform of -COOH groups

Lipoic Acid 1) Oxidizes acyl groups2) Acyl group transfer

Thiamine pyrophosphate

TPP

1) Formation of acyloins

(a-hydroxyketones)2) Oxidative decarboxylations ofa-ketoacids3) Non-oxidative decarboxylations


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COENZYMES - 3

While not strictly speaking coenzymes, these chemicals also

bring about changes.

1) Hydrolysis2) Hydration

1) Oxidations (hydroxylases) (oxidases)2) Oxidative couplings (phenols)

H2O

O2

Water

Oxygen

This list is not exhaustive, but it includes the majorcoenzymes that we will be discussing.

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