IM3'2 Part II Biosynthesis of Amino Acids and Nucleotides

1
Sources of NH3 for cells
Some bacteria can fix N2
Nitrogenase
N2
NH3
Plants and microbes can reduce nitrate
Nitrate reductase
Nitrite reductase
NO2-
NO3-
NH3
2
Assimilation of ammonia Bacteria incorporate NH3
into organic compounds in 2 ways 1. Reductive
amination of ?-ketoglutarate to form GLU. 2.
Amidation of the side-chain carboxyl groups of
GLU and ASP to form GLN and ASN.
3
Glutamate dehydrogenase catalyses formation of
a-ketoglutarate imine
and reduction to give glutamate.
4
Glutamine synthetase forms GLN from GLU and NH3
. This reaction requires ATP.
First the enzyme forms glutamate 5-phosphate, a
reactive intermediate.
5
Then NH3 displaces phosphate to form the amide.
ASN synthetase forms ASN from ASP and NH3 in the
same way.
6
Carboxyl group activation. An unactivated
carboxyl group is unreactive towards
nucleophilic attack. Carboxyl groups are often
activated by reaction with ATP to form acyl
phosphates.
7
Glutamate synthase catalyses the reaction GLN
?-ketoglutarate NADH H
GLU GLU NAD
The amide of GLN is hydrolysed, the NH3 released
is used for reductive amination of
a-ketoglutarate.
8
The GLN synthetase and glutamate synthase
combination is efficient.
GLU dehydrogenase is less efficient at
assimilating NH3
9
GLN is the source of amino groups in the
biosynthesis of several other compounds
purines, TRP, HIS and carbamoyl phosphate (a
precursor of pyrimidines and ARG).
The amino group of GLU can be transferred to
other ?-keto acids.
GLU ?-keto acid X
?-ketoglutarate amino acid X
Transaminases catalyse these group transfers.
10
Transaminases have pyridoxal phosphate (PLP) as
a cofactor.
Pyridoxal phosphate
Pyridoxamine
PLP-enzymes catalyse many different reactions of
amino acids.
11
The a-amino group of the AA condenses with the
aldehyde group of PLP to form an imine.
12
Any of the remaining 3 bonds may be cleaved so as
to convert the a-carbon to a carbanion.
The bond that is broken is orientated perpendicula
r to the plane of the pyridine ring.
Decarboxylases break the bond between Ca and the
carboxyl group.
Serine hydroxymethyltransferase cuts off the side
chain of serine.
13
Racemases and transaminases remove the a proton.
The pyridine ring acts as an electron sink that
effectively stabilises the negative charge.
14
Addition of a proton to the carbonyl carbon of
PLP gives a compound that is the imine of an
a-ketoacid and pyridoxamine.
15
Hydrolysis of the imine releases the a-ketoacid.
16
The sequence is then reversed to convert a
different a-ketoacid to an AA.
17
(No Transcript)
18
PLP-dependent enzymes can also catalyse
elimination and replacement reactions at the ?
and ? C atoms of AAs.