69. Dr. Shivani Pandey, Department of Biochemistry, K.G. Medical
University, Lucknow
Pyridoxine
70. PYRIDOXINE (vitamin B6)
Pyridoxal (PL) Pyridoxamine (PM)
Pyridoxine or
Pyridoxol (PN)
Each of these forms can be phosphorylated at position 5 to form: PLP, PMP & PNP
Dr. Shivani Pandey, Department of Biochemistry
71. Active form
Pyridoxal phosphate (PLP)
• PLP and PL account for 90% of the
total B6 in the blood.
• In the blood B6 is transported both
in the plasma and the RBCs.
• In the blood PLP is hydrolyzed to
PL because only free PL gets inside
the cells.
• In muscle and other tissues, PL is
converted back to PLP by a reversible
reaction with the help of alkaline
phosphatase and pyridoxal kinase.
• Active functional form is pyridoxal
phosphate (PLP) and pyridoxamine
phosphate (PMP).
• For absorption, the “phosphorylated”
form must be hydrolyzed to
“dephosphorylated” form by the enzyme
alkaline phosphatase in the intestine.
• In the portal vein Vit B6 is present as PL,
PM, PN.
• In the liver they are converted back to
phosphorylated forms. This conversion is
catalyzed by the ATP requiring enzyme,
pyridoxal kinase.
Dr. Shivani Pandey, Department of Biochemistry
72. Functions
FUNCTIONS: B6 is involved in:
Amino acid metabolism
Transamination reactions required for the synthesis and catabolism
of the amino acids.
Decarboxylation reactions.
Breakdown of glycogen Glycogenolysis (cofactor for glycogen phosphorylase).
80-90% of body vit B6 is present in the muscles, most of it in PLP
(coenzyme) form bound to glycogen phosphorylase. Only 1 mol or
less is present in the blood,
Synthesis of epinephrine (adrenaline) and norepinephrine (noradrenaline)
Synthesis of niacin (vitamin B3) from the amino acid tryptophan.
Dr. Shivani Pandey, Department of Biochemistry
73. Covalent bonds of -amino acids made labile by their
binding to PLP-containing enzyme
In the reactions of amino acid metabolism, the formyl (CHO) group of PLP
condenses with -NH2 group of an amino acid and forms a Schiffs base. This
linkage weakens or labilizes all the bounds around the -carbon of the amino acid.
The specific bond of an amino acid that is broken depends on the particular
enzyme to which PLP is attached.
Dr. Shivani Pandey, Department of Biochemistry
75. Deficiency
Food sources:
In animal foods major forms are PL and and PM along with their phosphorylated forms.
In plants PN.
Bananas, beans, lentils, walnuts, salmon, chicken, beef, whole grain breads and cereals,
soybeans, liver, eggs, dairy products are excellent sources.
Requirements:
The requirement for vitamin B6 in the diet is proportional to the level of protein
consumption ranging from 1.4 - 2.0 mg/day for a normal adult.
During pregnancy and lactation the requirement for vitamin B6 increases
approximately 0.6 mg/day.
TOXICITIES:
Megadoses of B6 (daily doses of >500mg) are used to treat pms symptoms. They can
cause neurotoxicity and photosensitivity in some individuals.
Deficiencies: are rare and usually are related to an overall deficiency of all the
B-complex vitamins.
Certain drugs form complexes with PL and PLP
Penicillamine (used to treat rheumatoid arthritis and cystinurias).
Isoniazid (the hydrazide derivative of isonicotinic acid) is the primary drug for
chemotherapy of tuberculosis.
Dr. Shivani Pandey, Department of Biochemistry
77. BIOTIN
Biotin is relatively small, bicyclic (two-ring) compound
formed from a tetrahydrothiophene (thiophene) ring ,
and a second ring, which contains a ureido group.
The thiophene ring also has a valeric acid side chain.
Although eight different stereoisomers of biotin exist, only
one stereoisomer is found naturally and to have biological
activity as a coenzyme. It is called d-(+)-biotin, D-biotin or
simply biotin.
It is
sometimes
called vitamin
H and also
coenzyme R.
Dr. Shivani Pandey, Department of Biochemistry
78. Holocarboxylase
In humans, the four holocarboxylases are : acetyl-CoA carboxylase,
propionyl-CoA carboxylase, pyruvate carboxylase and beta-
methylcrotonyl-CoA carboxylase. Biotin is chemically bonded in each of these
enzymes via an amide linkage between the carboxyl group of the valeric acid
side-chain in biotin and the epsilon-amino group of the lysine residue in the
apocarboxylase.
The enzyme that catalyzes the formation of this covalent bond is called
holocarboxylase synthetase. Dr. Shivani Pandey, Department of Biochemistry
79. Biotin Cycle
Biotin cycle: the chain of chemical reactions involved in the use and reuse of the
vitamin biotin. One important role of biotinidase is:
1. To separate or free biotin from proteins to which it is bound in foods. Biotin in its
free form can then be used by the body.
2. Biotinidase lets the body recycle or reuse the biotin over and over again so that we
do not need to consume large amounts of this vitamin in our diets.
•Within cells, the carboxylases
(pyruvate carboxylase, acetyl-CoA
carboxylase, methycrotonyl-CoA
carboxylase, propionyl-CoA
carboxylase) are biotinylated via
holocarboxylase synthetase. Biotin
and apocarboxylases are the
substrates. ATP and magnesium also
participate in the reaction.
Biotinidase deficiency is a treatable,
inherited metabolic disorder in which the
body cannot process the vitamin biotin in a
normal manner.
Dr. Shivani Pandey, Department of Biochemistry
80. Functions
Coenzyme involving CO2 transfer
acetyl CoA to malonyl CoA
Propionyl CoA to methylmalonyl CoA
Pyruvate to oxaloacetate
Deficiency
Fatty liver and kidney syndrome, a fatal metabolic
disorder in chicks and turkey poults
Raw egg white (avidin) can block absorption
Dr. Shivani Pandey, Department of Biochemistry
81. Biotin (functions)
Coenzyme for several reactions involving CO2 fixation into various
compounds e.g.
Pyruvate to oxaloacetate
(pyruvate carboxylase)
Propionyl CoA to methylmalonyl CoA
(propionyl CoA carboxylase)
Acetyl CoA to malonyl CoA
(acetyl CoA carboxylase)
- initial step in de novo fatty
acid synthesis.
Dr. Shivani Pandey, Department of Biochemistry
82. Deficiency symptoms
Rare because of widespread distribution in plant and
animal food and significant lower gut synthesis.
Sources
Yeast, rice, soybeans, peanuts, fish (herring and
mackerel), mushrooms and bananas, safflower meal,
liver and milk are rich sources.
Can be induced by eating raw egg white
The fact is that nature created the egg in such a way that
its yolk is very rich in biotin. One of the highest
concentration in nature. Eat the egg whole together with the
egg white and you will be fine.
Egg whites contain a glycoprotein called "avidin" which
binds biotin - one of the B vitamins - very effectively. The
cooking process deactivates the avidin in the egg, much the
same way it deactivates every other protein in the egg
white.
Biotin deficiency is chief cause of fatty liver and kidney
syndrome.
This baby developed
severe biotin deficiency
during intravenous feeding
without biotin.
Aajonus Vonderplanitz,
in his book “We Want to
live” is a strong proponent
of raw eggs.
Dr. Shivani Pandey, Department of Biochemistry
83. How Biotin Works
1- Biotin carrier protein
2- Biotin carboxylase
3- Transcarboxylase
Dr. Shivani Pandey, Department of Biochemistry
85. FOLIC ACID (folacin)
• Folacin includes several derivatives of folic acid
(monopteroylglutamic acid).
• Folic acid is obtained primarily from yeasts and leafy vegetables as
well as animal liver. Animals cannot synthesize PABA nor attach
glutamate residues to pteroic acid, thus, requiring folate intake in the
diet.
“Microorganisms
Only can
synthesize Folacin”
Sulphonamides
and antibiotics
block PABA
incorporation
by competitive
inhibition
Dr. Shivani Pandey, Department of Biochemistry
86. Structure
Folic acid exists in a polyglutamate form. Intestinal mucosal cells
remove some of the glutamate residues through the action of the
lysosomal enzyme, conjugase.
Dr. Shivani Pandey, Department of Biochemistry
87. Humans and microorganisms:
Folic acid is reduced within cells
(principally in the liver where it is
stored) to tetrahydrofolate (THF or
H4folate) through the action of
folate reductase [or
dihydrofolate reductase (DHFR) ]
which is an NADPH-requiring
enzyme.
Active functional form is:
Tetrahydrafolic acid
(THF).
Dr. Shivani Pandey, Department of Biochemistry
88. Active center (N5 and N10)
Dr. Shivani Pandey, Department of Biochemistry
89. • Active center of tetrahydrofolate (THF). The N5 position is the site of
attachment of methyl and formimino groups, the N10 the site for
attachment of formyl group and that both N5 and N10 bridge the
methylene and methenyl groups.
Dr. Shivani Pandey, Department of Biochemistry
90. Folate conversions
Carrier of one-carbon (e.g. methyl) groups
The one-carbon units are added to, or removed from, metabolites such as:
• histidine
• serine
• methionine
• purines.
Dr. Shivani Pandey, Department of Biochemistry
91. Functions
• Role of N5,N10-methylene-THF in dTMP
synthesis is the most metabolically
significant function for this vitamin.
• Vitamin B12 and N5-methyl-THF in the
conversion of homocysteine to
methionine is important in helping cells to
regenerate needed THF.
Dr. Shivani Pandey, Department of Biochemistry
92. Participation of H4folate in dTMP synthesis
______Deoxyuridine______________ ________Deoxythymidine
____Monophosphate (dUMP)_______________Monophosphate (dTMP)_______
Dr. Shivani Pandey, Department of Biochemistry
94. Deficiency symptoms
• Identical to those for vitamin B12 deficiency:
• Effect of folate deficiency on cellular processes
is upon DNA synthesis.
– Impairment in dTMP synthesis and purine synthesis
– Cell cycle arrest in S-phase of rapidly proliferating cells, especially
hematopoietic cells.
• The result is megaloblastic leukemia as for vitamin B12 deficiency.
– The inability to synthesize DNA during erythrocyte maturation leads to
abnormally large erythrocytes termed macrocytic anemia.
• Deficiency during pregnancy can cause neural tube defects such as
spina bifidia.
Deficiency is rare due to the adequate presence of folate in food.
Deficiency can occur when there is:
1. Poor dietary habits as those of chronic alcoholics.
2. Impaired absorption or metabolism
• Certain drugs such as anticonvulsants and oral contraceptives and
methotrxate can impair the absorption of folate.
3. An increased demand for the vitamin.
• Pregnancy
• Folate will nearly double by the third trimester of pregnancy.
Dr. Shivani Pandey, Department of Biochemistry
95. What are neural tube defects?
• neural tube defects (NTDs) are birth defects
that occur early in pregnancy
– often before a woman knows she is pregnant
• neural tube grows to become baby’s spinal
cord, spine, brain and skull
Dr. Shivani Pandey, Department of Biochemistry
96. The neural tube forms in the embryo and
then closes (between the 2nd and 4th week of
gestation)
A neural tube defect occurs when the neural
tube fails to close properly
The two most common NTDs are
anencephaly and spina bifida
Dr. Shivani Pandey, Department of Biochemistry
98. Which NTDS are most
common?
• Spina Bifida
– a condition that results when
the lower part of the neural
tube fails to develop properly
• Anencephaly
– a fatal condition in which the
upper end of the neural tube
fails to close
Occult
a
Meningocele
Myelomeningocele
Dr. Shivani Pandey, Department of Biochemistry
100. How Can NTDs be Prevented?
• All women of childbearing age should receive 0.4
mg (400 micrograms) of folic acid daily prior to
conception of planned or unplanned pregnancies
and continue thru 1st trimester
• Women with a history of NTD and should receive
daily supplementation of 4 mg (4000 micrograms)
of folic acid starting three months prior to
conception and continuing thru the 1st trimester
Dr. Shivani Pandey, Department of Biochemistry
102. VITAMIN B12 (cobalamin)
• Vitamin B12, is also called cobalamin,
cyanocobalamin and hydroxycobalamin.
• It is built from :
1. A nucleotide and
2. A complex tetrapyrrol ring structure
(corrin ring)
3. A cobalt ion in the center.
4. A R- group
• When R is cyanide (CN), vitamin B12
takes the form of cyanocobalamin.
• In hydroxycobalamin, R equals the
hydroxyl group (-OH).
• In the coenzyme forms of vitamin B12,
– R equals an adenosyl group in
adenosylcobalamin.
– R equals a methyl (-CH3) group in
methylcobalamin.
• Vitamin B12 is synthesized exclusively by
microorganisms (bacteria, fungi and algae)
and not by animals and is found in the liver
of animals bound to protein as
methycobalamin or 5'-
deoxyadenosylcobalamin.
Dr. Shivani Pandey, Department of Biochemistry
103. • Known as the "red" vitamin because it
exists as a dark red crystalline compound,
Vitamin B12 is unique in that it is the only
vitamin to contain cobalt (Co3+) metal ion,
which, gives it the red color.
• The vitamin must be hydrolyzed from
protein in order to be active.
• Intrinsic factor, a protein secreted by
parietal cells of the stomach, carries it to the
ileum where it is absorbed.
• It is transported to the liver and other
tissues in the blood bound to
transcobalamin II.
• It is stored in the liver attached to
transcobalamin I.
– It is released into the cell as
Hydroxocobalamin (see the next slide)
• In the cytosol it is converted to
methylcobalamin.
• Or it can enter mitochondria and be
converted to 5’-deoxyadenosyl
cobalamin.
Dorothy Crowfoot Hodgkin
(1910-1994)
Dr. Stadtman in her lab
Dr. Shivani Pandey, Department of Biochemistry
104. In the cytosol
In mitochondria
Dr. Shivani Pandey, Department of Biochemistry
105. Functions
• Only two reactions in the body require vitamin B12 as a
cofactor:
1. During the catabolism of fatty acids with an odd number of
carbon atoms and the amino acids valine, isoleucine and
threonine the resultant propionyl-CoA is converted to
succinyl-CoA for oxidation in the TCA cycle.
– methylmalonyl-CoA mutase, requires vitamin B12 as a cofactor in
the conversion of methylmalonyl-CoA to succinyl-CoA.
– 5'-deoxyadenosine derivative of cobalamin is required for this
reaction
2. The second reaction catalyzed by methionine synthase
converts homocysteine to methionine
– This reaction results in the transfer of the methyl group from N5-
methyltetrahydrofolate to hydroxycobalamin generating
tetrahydrofolate and methylcobalamin during the process of the
conversion.
Dr. Shivani Pandey, Department of Biochemistry
107. Deficiency symptoms
• Pernicious anemia in humans (inability to absorb B12
because of lack of gastric intrinsic factor).
• Neurological disorders due to progressive demyelination
of nerve cells.
– This results from increase in methylmalonyl-CoA.
– Methylmalonyl-CoA is a competitive inhibitor of malonyl-CoA in
fatty acid biosynthesis.
– Can substitute malonyl-CoA in any fatty acid biosynthesis and
create branched-chain fatty acid altering the architecture of
normal membrane structure of nerve cells.
• Sources
– Synthesized only by microorganisms, so traces only are present
in plants; liver is a rich source.
– B12 is found in organ and muscle meats, fish, shellfish, dairy
products, eggs and in fortified foods like breakfast cereals.
Dr. Shivani Pandey, Department of Biochemistry