1. Dr. Ketki K Assistant Professor Dept of Biochemistry HIMS Varanasi
Nitric oxide(NO), Polyamies, Biogenic amines & Branched Chain Amino Acid Metabolism
Dr. Ketki K Assistant Professor Dept of Biochemistry
HIMS Varanasi

2. Nitric Oxide:
Chemistry
Metabolic fate
Isoenzymes of NOS
MOA
Physiological action
Diseases & treatment

3. Nitric Oxide Chemistry: Highly reactive “free radical”
Half life : 3-4 second
Nitric oxide synthase (NOS) reaction
contains heme, FAD, FMN, NADPH & tetrahydrobiopterine
Calmodulin modulates its activity
The guanidino nitrogen of arginine is incorporated
into NO'
From the molecular oxygen, one atom is added to
NO'and the other into citrulline
Therefore, the enzyme is a di-oxygenase

5. Metabolic fate Nitric oxide : short half life (3-4 seconds)
NO + O2 = NO2 → excreted in urine
Very low quantity of NO: expelled through lung
On exposure to superoxide anion (O2‘)
nitric oxide (NO' ) is converted to a highly reactive free radical, peroxy nitrite(OONO'), which causes lipid peroxidation, cell injury and cell death

6. Iso-enzymes of NOS Neuronal NOS Macrophage NOS Endothelial NOS
Other name
nNOS
iNOS (inducible)
eNOS
Site
Central & peripheral neurons (cerebellum,GIT)
Macrophages,
neutrophils,
hepatocytes
Endothelial cells,
Platlets
Endocardium
myocardium
Location
Cytoplasmic
Plasma membrane
Effect of calcium
Activates it
Does not activate it,induced by cytokines &during inflammation
Gene
Chromosome 12
Chromosome 17
Chromosome 7

7. MOA of NO
In smooth muscles, NO activates guanylate cyclase ↓ Increased cGMP Activates protein kinase dephosphorylation of myosin light chain Relaxation of smooth muscles Vasodilatation

8. Physiological actions of NO
Blood vessels:
endothelial NOS (NOSe) present
Potent vasodilator
Deficiency of NO is a/w hypertension
Excessive production of NO results in refractory hypotension, seen in patients with septicemic shock

9. 2) In CNS, NOSn isoform is present
2) In CNS, NOSn isoform is present. Glutamate acts on NMDA receptors: longstanding calcium influx This activates NOSn. NO‘ stimulates the releasing hormones (CRH, GHRH and LHRH) Acts as neurotransmitter

10. 3) Macrophages: isoform NOSi (i stands for inducible)
3) Macrophages: isoform NOSi (i stands for inducible). peroxy nitrite: lethal to micro-organisms(bactericidal) 4) Platelets: inhibits adhesion of platelets 5) Intestine: relaxes smooth muscles of GIT, reduces GI motility

11. Nitric oxide in diseases & treatment
Angina pectoris: Nitroglycerin
Pulmonary Hypertension: inhalation of NO
Impotence :Sildenafil: PDE-5 inhibitor

12. Polyamines Having multiple amino groups
Putrescine, spermidine and spermine
(last 2 detected in human sperm/semen)
Key enzyme of synthesis: ODC
(short half life: 10 mt)
requires PLP
induced by steroid hormones

13. Biosynthesis
SAM decarboxylase: Pyruvate as prosthetic group, only mamallian enzyme known to have bound pyruvate as prosthetic group

14. Degradation
Excreted in urine, as conjugated form, as acetylated derivatives
Oxidized to ammonia & CO2

15. Functions Involved in synthesis of DNA/RNA/Protein
Essential for cell growth & proliferation
Stabilizes membrane structure
Protein kinases inhibited by polyamines

16. Clinical importance
DFMO(difluromethyl ornithine) : inhibits polyamine synthesis in parasites
useful against trypanosomes of Indian Kala azar & pneumocystis carnii parasite in AIDS
2) Excretion of polyamines : elevated in urine in all cancers eg: leukemia, ca lung,kidney ,bladder

17. Biogenic amines
Decarboxylation of amino acids or their derivatives : forms biogenic amines

19. Branched chain amino acid metabolism
Branched chain amino acids
Valine(V): glucogenic,
Leucine(L): Ketogenic
Isoleucine(I): both glucogenic & ketogenic
All are essential aa
Provide fuel for brain during starvation

21. Second enzyme of the pathway:
branched chain alpha keto acid dehydrogenase
Complex of decarboxylase, transacylase and dihydrolipoyl dehydrogenase
So, this resembles pyruvate dehydrogenase

22. branched chain alpha keto acid dehydrogenase
Complex mitochondrial enzyme
Oxidative decarboxylation
5 co-enzymes: TPP,Lipoamide,FAD,Coenzyme A,NAD+

23. Metabolic defects Maple syrup urine disease
Intermittent branched chain ketonuria
Isovaleric acidemia
Hypervalinemia

24. Maple syrup urine disease
Also c/a branched chain ketonuria
Incidence: 1 in 1 lakh births
Name originates from the characteristic smell of urine (similar to burnt sugar or maple sugar)due to excretion of branched chain keto acids

25. Basic biochemical defect :
Deficient decarboxylationof branched chain keto acids(BKA) (reaction 2)

27. Clinical findings:
Disease starts in the first week of life.
characterized by
Convulsions, severe mental retardation, vomiting, acidosis, coma
death within the first year of life

28. Laboratory findings:
Urine contains branched chain keto acids,valine, leucine and isoleucine.
Rothera's test is positive
Diagnosis depends on enzyme analysis in cells
Preferably within first week of life

29. Treatment:
Giving a diet low in branched chain amino acids
Mild variant is called intermittent branched chain ketonuria
This will respond to high doses of thiamine. This
is because the decarboxylation requires thiamine
Liver transplantation has been successfully tried in some cases of MSUD

30. Isovaleric acidemia:
Metabolic defect: deficiency of isovaleryl CoA dehydrogenase
Consequences: increased isovalerate in urine
C/F: Mild Mental retardation, “cheesy” odor in
breath & body fluids

32. Hypervalinemia:
Metabolic defect: deficiency of enzyme responsible for transamination of valine alone
Consequences: increased valine in plasma

35. Homework Aminoacids & their derivatives as neurotransmitters
Table 15.7 satyanarayan