good

1. NON PROTEIN
AMINOACIDS

2. Non protein amino acids
 These amino acids, although never found
in proteins, perform several biological
important function.
 These NPAAs and D-AA speculated to be
related to auto immune disease and to
aging

3. understand the role of NPAAs and D-
AA s in auto immune disease and
aging, the determination of these
NPAAs and D-AA s is required.
 any nonprotein amino acid can be
chemically incorporated into peptides,
provided that appropriate methods are
designed for protecting the functional
group.

4.  Nonprotein amino acids with no
cytotoxicity have been known to be
incorporated into proteins. For examples,
 tyrosine and tryptophan residues in some
proteins have been substituted with m-
fluorotyrosine and 4-fluorotryptophan
respectively, without any effects on the
protein functions, and the 19F nuclei have
been used as magnetic resonance

5. One NPA that has received some attention is canavanine,
(L-2-amino-4-(guanidinooxy)butyric acid), the
guanidinooxy structural analogue of arginine.
These non protien amino acids are classified as alpha
and non alpha amino acids
1) Alpha amino acids:
a) ornithine
b) citruline
c) arginosuccinic acid
d) thyroxine
e) triodothyroxine
f) S-Adenosylmethionine
g) Homocysteine
h) 3,4-Dihydroxy phenylalanine ( DOPA)
I ) creatinine
j) ovathiol
k) Azaserine

6.  2) NON ALPHA amino acids:
a) beta –alanine
b) beta – aminoisobutyric acid
c) gama – aminobutyric acid(GABA)
d) aminolevulinic acid (ALA)
e) taurine

7. Alpha - aminoacids
1) ornithine :
 ornithine is precursors of polyamine
Hydrolytic cleavage of the guanidino group of
arginine, catalyzed by liver arginase, releases urea .
The other product, ornithine, reenters liver
mitochondria and participates in additional rounds of
urea synthesis.
Ornithine and lysine are potent inhibitors of arginase,
and compete with arginine.
Arginine also serves as the precursor of the potent
muscle relaxant nitric oxide (NO) in a Ca2+-
dependent reaction catalyzed by NO synthase
2) Citrulline :
Citrulline is intermediates in the biosynthesis of urea

8. 
L-Ornithine transcarbamoylase catalyzes
transfer of the carbamoyl group of carbamoyl
phosphate to ornithine,forming citrulline &
orthophosphate While the reaction occurs in
the mitochondrial matrix, both the formation of
ornithine and the subsequent metabolism of
citrulline take place in the cytosol.
 Entry of ornithine into mitochondria and
exodus of citrulline from mitochondria
therefore involve mitochondrial inner
membrane transport systems

9. Arginosuccinic acid :
 Arginosuccinic acid is intermediates in
the biosynthesis of urea
 Argininosuccinate synthetase links
aspartate and citrulline via the amino group
of aspartate and provides the second
nitrogen of urea.
 The reaction requires ATP and involves
intermediate formation of citrullyl-AMP.
Subsequent displacement of AMP by
aspartate then forms arginosuccinate.

10. In addition to patients that lack
detectable argininosuccinate
synthetase activity a 25-fold elevated
Km for citrulline has been reported. In
the resulting citrullinemia, plasma and
cerebrospinal fluid citrulline levels are
elevated, and 1–2 g of citrulline are
excreted daily.

11. 4) Thyrosine and triodothyroxine
:
 Tyrosine forms norepinephrine and
epinephrine, and following iodination the
thyroid hormones triiodothyronine and
thyroxine.
 Use of measurement of blood thyroxine
or thyroid-stimulating hormone (TSH) in
the neonatal diagnosis of congenital
hypothyroidism.
 The amino acid tyrosine is the starting
point in the synthesis of the
catecholamines and of the thyroid
hormones tetraiodothyronine (thyroxine;
T4 ) and triiodothyronine (T3)

12.  thioredoxin reductase, glutathione
peroxidase, and the deiodinase that converts
thyroxine to triiodothyronine.
 The clinical history, physical examination,
and lab results were all consistent with
primary hypothyroidism. Accordingly, the
patient was started on a low dose of
thyroxine (T4 ).
 It is important to begin therapy with a small
dose of T4, as larger doses can precipitate
serious cardiac events, due to the changes in
metabolism caused by administration of the
hormone.
 Thyroxine (T4), free: 4.0 pmol/L (normal
10.3–21.9 pmol/L)

13. 5)S-Adenosylmethinine
homocysteine:S-Adenosylmethionine, the principal source of methyl groups
in metabolism, contributes its carbon skeleton to the
biosynthesis of the polyamines spermine and spermidine.
Homocystinuria
Homocystinuria Cystathionine -synthase Lens dislocation,
thrombotic vascular disease, mental retardation, osteoporosis
AR
Homocystinuria
5,10-Methylenetetrahydrofolate reductase
Mental retardation, gait and psychiatric abnormalities,
recurrent strokes ,Mental retardation, hypotonia, seizures,
megaloblastic anemia

14.  Pathways, enzymes, and coenzymes involved
in the homocystinurias. Methionine transfers a
methyl group during its conversion to
homocysteine.
 Defects in methyl transfer or in the subsequent
metabolism of homocysteine by the pyridoxal
phosphate (vitamin B6)-dependent cystathionine
b-synthase increase plasma methionine levels.
 Homocysteine is transformed into methionine via
remethylation. This occurs through methionine
synthase, a reaction requiring methylcobalamin
and folic acid.
 Deficiencies in these enzymes or lack of cofactors
is associated with decreased or normal methionine
levels. In an alternative pathway, homocysteine
can be remethylated by betaine:homocysteine
methyl transferase

15.  Life-threatening vascular complications
(affecting coronary, renal, and cerebral
arteries) can occur during the first decade of
life and are the major cause of morbidity and
mortality.
 Classic homocystinuria can be diagnosed
with analysis of plasma amino acids,
showing elevated methionine and presence
of free homocystine.

16.  Total plasma homocysteine is also extremely
elevated (usually >100 M). Treatment consists
of a special diet restricted in protein and
methionine and supplemented with cystine.
 In approximately half of patients, oral
pyridoxine (25–500 mg/d) produces a decrease
in plasma methionine and homocystine
concentration in body fluids.
 Folate and vitamin B12 deficiency should be
prevented by adequate supplementation.
Betaine is also effective in reducing
homocystine levels.

18. 6) 3-4
Dihydrophenylalanine(DOPA):
 Neural cells convert tyrosine to epinephrine and
norepinephrine. While dopa is also an intermediate in
the formation of melanin, different enzymes hydroxylate
tyrosine in melanocytes.
 Dopa decarboxylase, a pyridoxal phosphate-dependent
enzyme, forms dopamine. Subsequent hydroxylation by
dopamine -oxidase then forms norepinephrine.
 In the adrenal medulla, phenylethanolamine-N-
methyltransferase utilizes S-adenosylmethionine to
methylate the primary amine of norepinephrine, forming
epinephrine .
 Tyrosine is also a precursor of triiodothyronine and
thyroxine.
 DOPA ... related to dopaminerelationship to Parkinson's
Disease

19.  Dopamine, Norepinephrine, and Epinephrine
1. SYNTHESIS OF THE CATECHOLAMINE
NEUROTRANSMITTERS
 These three neurotransmitters are synthesized in a
common pathway from the amino acid L-tyrosine.
 The first and rate-limiting step in the synthesis of these
neurotransmitters from tyrosine is the hydroxylation of
the tyrosine ring by tyrosine hydroxylase, a
tetrahydrobiopterin(BH4)-requiring enzyme. The
product formed is dihydroxyphenylalanine or DOPA.
 The phenyl ring with two adjacent OH groups is a
catechol, andhence dopamine, norepinephrine, and
epinephrine are called catecholamines.

20.  The second step in catecholamine synthesis is the
decarboxylation of DOPA to form dopamine. This
reaction, like many decarboxylation reactions of
amino acids,equires pyridoxal phosphate.
 Dopaminergic neurons (neurons using dopamine
as a neurotransmitter) stop the synthesis at this
point, because these neurons do not synthesize
 the enzymes required for the subsequent steps.
Neurons that secrete norepinephrine synthesize it
from dopamine in a hydroxylation reaction
catalyzed by dopamine -hydroxylase (DBH). This
enzyme is present only within the storage vesicles
of these cell
 Although the adrenal medulla is the major site of
epinephrine synthesis, it is also synthesized in a
few neurons that use epinephrine as a

22. 7)Creatinine :
Creatinine is formed in muscle from creatine
phosphate by irreversible, nonenzymatic
dehydration and loss of phosphate.
Since the 24-h urinary excretion of creatinine is
proportionate to muscle mass, it provides a
measure of whether a complete 24-h urine
specimen has been collected.
 Glycine, arginine, and methionine all participate
in creatine biosynthesis. Synthesis of creatine
is completed by methylation of guanidoacetate
by S-adenosylmethionine.

24. Normal values :
 Creatinine : 200 mol/L (44–80 mol/L)
 Female -- 44–80 mol/L 0.5–0.9
ng/mL
 male -- 53–106 mol/L 0.6–1.2
ng/mL

25. 8) Ovathiol:-
 Sulfur containing amino acid found in fertilized
Eggs, and acts as an antioxidant
9) Azaserine: (antibiotic)
Purine deficiency states, while rare in humans,
generally reflect a deficiency of folic acid.
Compounds that inhibit formation of
tetrahydrofolates and therefore block purine
synthesis have been used in cancer
chemotherapy.
Inhibitory compounds and the reactions they inhibit
include azaserine, diazanorleucine, 6-
mercaptopurine , and mycophenolic acid .

26. II) Non--Amino Acids:
 Non--amino acids present in tissues in a free form
include -alanine, -aminoisobutyrate, and -aminobutyrate
(GABA). -Alanine is also present in combined form in
coenzyme A and in the -alanyl dipeptides carnosine,
anserine and homocarnosine .
 1) Beta-Alanine & -Aminoisobutyrate :
 Alanine and -aminoisobutyrate are formed during
catabolism of the pyrimidines uracil and thymine,
respectively . Traces of -alanine also result from the
hydrolysis of -alanyl dipeptides by the enzyme
carnosinase. -Aminoisobutyrate also arises by
transamination of methylmalonate semialdehyde, a
catabolite of L-valine .
 The initial reaction of -alanine catabolism is
transamination to malonate semialdehyde. Subsequent
transfer of coenzyme A from succinyl-CoA forms
malonyl-CoA semialdehyde, which is then oxidized to
malonyl-CoA and decarboxylated to the amphibolic
intermediate acetyl-CoA.

27. Analogous reactions characterize the catabolism
of -aminoisobutyrate. Transamination forms
methylmalonate semialdehyde, which is converted
to the amphibolic intermediate succinyl-CoA by
reactions 8V and 9V of.
Disorders of -alanine and -aminoisobutyrate
metabolism arise from defects in enzymes of the
pyrimidine catabolic pathway.
Principal among these are disorders that result
from a total or partial deficiency of
dihydropyrimidine dehydrogenase.

28. 2) beta-Alanyl Dipeptides :
 The -alanyl dipeptides carnosine and anserine
(N -methylcarnosine) activate myosin ATPase,
chelate copper, and enhance copper uptake. -
Alanyl-imidazole buffers the pH of anaerobically
contracting skeletal muscle.
 Biosynthesis of carnosine is catalyzed by
carnosine synthetase in a two-stage reaction tha
involves initial formation of an enzyme-bound
acyl-adenylate of -alanine and subsequent
transfer of the -alanyl moiety to L-histidine.

29.  Hydrolysis of carnosine to -alanine and L -
histidine is catalyzed by carnosinase. The
heritable disorder carnosinase deficiency is
characterized by carnosinuria.
 Homocarnosine, present in human brain at
higher levels than carnosine, is synthesized in
brain tissue by carnosine synthetase. Serum
carnosinase does not hydrolyze
homocarnosine. Homocarnosinosis, a rare
genetic disorder, is associated with progressive
spastic paraplegia and mental retardation.

30. 3) gama-Aminobutyrate
 gama-Aminobutyrate (GABA) functions in brain
tissue as an inhibitory neurotransmitter by
altering transmembrane potential differences.
 GABA is formed by decarboxylation of
glutamate by L -glutamate decarboxylase.
Transamination of -aminobutyrate forms
succinate semialdehyde, which can be reduced
to -hydroxybutyrate by L -lactate
dehydrogenase, or be oxidized to succinate
and thence via the citric acid cycle to CO2 and

31.  A rare genetic disorder of GABA metabolism
involves a defective GABA aminotransferase,
an enzyme that participates in the catabolism of
GABA subsequent to its postsynaptic release in
brain tissue.
 Defects in succinic semialdehyde
dehydrogenase are responsible for another rare
metabolic disorder of -aminobutyrate
catabolism characterized by 4-hydroxybutyric
aciduria.

33.  4) amino levulinic acid (ALA) :
 ALA is intermediate in the synthesis of
porphyrin (finally heme)

34. 5) Taurine :
 Taurine (2-aminoethylsulphonic acid) is a non-
protein
aminoacid present in almost all animal tissues and
the most abundant free intracellular aminoacid in
human cells.
 In humans, it is considered to be a “semi-essential
aminoacid” since it can be synthesized from
other sulfonic aminoacids such as methionine and
cysteine, in
the presence of vitamin B6,2,3 but endogenous
production is insufficient, so that it needs to be
provided through diet.

36.  Biological effects of taurine in the context of
diabetes
 Biological EffectMechanismof Taurine
 Antioxidant action By inhibiting ROS generation
at mitochondria Osmoregulation By
counteracting osmotic inbalance through
cellular membrane due
 to hyperglycaemia
 Antiinflammatory effects By interfering the
formation of inflammatory mediators Glucose
Homeostasis By interfering the insulin
signalling pathway acting upon UCP2 protein

37.  Estimation of NPAAs :
1) The aim of our study was to analyze NPAAs
and D-AA s in biosamples by means of capillary
electrochromatogrphy (CEC) using a chiral
practicle- loded monolithiac column with
flurrosense detection for high sensitivity.
2) capillary electrophoresis
3) High perfromance liquid
chromatography(HPLC)
4) laser-induced flurosecne (LIF)
5) scanning electro microscopy (SEM)

38.  Scanning electro micrograph of CEC
capillary coulmn

39. Location of the regions of ordered secondary structures for b-residues in
f–q–y space. The a-helix and b-sheet are the classical structures for poly a-
amino
acids. b-residues occurring in the appropriate shaded region can be

40. THANK Q