Download Biochemistry –Second year, Coll

Biochemistry –Second year, Coll. of Medicine-Baghdad Univ.
2011-2012.
Dr.Basil Oied Mohammed Saleh.
Subjec:Lipid
Lecture 5,6,7
Objective: To illustrate the steps involved in production of energy ATP from
oxidation of fatty acids .
The fate of the saturated Fatty Acids
The synthesized saturated fatty acids, mainly in the liver and those derived
from the diet as chylomicrone as mentioned ,Lecture 2 are stored as
triglyceride(triacylglycerol) in adipose tissues and represents the principal
energy reserve in human body. TG which referred to as fat when its solid and
as oil when it is liquid at room temperature is not normally stored in the liver
as fatty liver is the early stage in different forms of chronic liver diseases;
alcoholic liver cirrhosis. The synthesis of TG occurred in the liver and adipose
tissues. In the steps of TG synthesis pathway, the synthetic units are; glycerol
and fatty acids which firstly must be activated into active forms to be ready for
their reaction. Glycerol is converted into glycerol phosphate(active form) which
in the liver is achieved by Glycerol kinase enzyme, the enzyme present only in
the liver
Glycerol kinase
Gylcerol …………………………………….» Glycerol phosphate
ATP……ADP+Pi
The formation of activated glycerol; glycerolphosphate is also activated by
another enzyme; the Glycerol phosphate dehydrogenase(GlyPDH) but in
different reaction:
Glycolysis pathway
GlyPDH
Glucose………………………..» DHAP………..……….» Glycerol phosphate
NADH….NAD +
This latter reaction occurred in liver and adipose tissues because the
activated enzyme is found in both these two organs.
Note: The adipocytes can take glucose from the circulation only in the
presence of Insulin, so in the case of fasting state or pathological cases as
in DM when the blood glucose and so the Insulin levels are normally low,
the adipocytes uptake of glucose and the synthesis of glycerol will be
limited with consequent inhibition of TG pathway synthesis. So, the
pathway of fatty acids and TG synthesis is stimulated in
the presence of Insulin Hormone and inhibited in its
absence.
The second substance in the synthesis of fatty acid, which activated by its
conversion into
acyl-CoA , the active form of fatty acid the reaction is
stimulated by the enzyme Thiokinases(or Fatty acyl CoA synthetases).
Thiokinases
RCOOH………………………………↑…….». RCO CoA
ATP….»AMP+PPi CoA
The activated two substances; Glycerol phosphate and Acyl CoA reacted in
four steps to form TG:
(1)
Acyltransferase
Glycerol phosphate+ Acyl CoA…↓….» Monoacylglycerol(Lysophosphatidic
acid)
CoA
↓
(2)
Acyltransferase ↓← AcylCoA
↓
Diacylglycerol(phosphatidic acid )
↓
H2O→ ↓→ Pi (3)
↓
Diacylgylcerol(Diglyceride)
(4)
Acyltransferase
Diglyceride………………………………»TG
↑
AcylCoA
The TG is stored in the adipose tissues as anhydrous form(insoluble in water)
and occupied the major space of adipocytes cytoplasm and is ready for
mobilization to be used for production of energy when there is need.
Mobilization and Oxidation of Fat
The stored TG in adipose tissues is released in the form of saturated fatty acids
and glycerol when there is need for the stored energy;prolonged fastin g,
starvation and DM. This process is referred to as LIPOLYSIS, which is the
opposite to lipogenesis. The Lipolysis is stimulated in the presence of the Insulin
antagonist, principally the Glucagon, adrenaline, ACTH, cortisol,TSH and
noradrenaline. Insulin is the potent inhibitor of lipolysis pathway.The key
enzyme in the lipolysis is: Hormone sensitive -lipoprotein Lipase(HS-LPL), the
enzyme which act only under stimulation of the insulin anagonist hormones.
These hormones by their binding to their CM receptors increased the synthesis
of the intracellular concentration of the second messanger
c AMP, which
inturn converts the HS-LPL from its inactive form into active form:
cAMP+PPi
.
↑
Hormones………………»CM(Receptors).
ATP this Hs+Receptors binding
Reaction leads to increase
Adrenaline,glucagon….
the cAMP intracellular
levels
The formed intracellular cAMP stimulates the formation of active protein
kinase which is important in conversion of HS-LPL into its active form:
Active Protein Kinase
HS-LPL( dephosphorylste form)………↑…………» HS-LPL (phosphorlyated)
( Inactive form)
cAMP
(Active form)
↑
.
Hs
Now:
HS-LPL
TG…………………↓………………..» Diacylglycerol in adipocytes…..»
Fatty acids
↓→fatty
These fatty acids are
HIS-LPL ↓
acids
Able to penetrate the CM
Monoacylglycerol
Of adipocytes and released
↓ fatty
Into the circulation.
.
↓ acids
Glycerol
The formed saturated Fatty acids in these three steps and the glycerol in the
last step are released into the circulation. The last two enzymes HIS-LPL
hormone insensitive LPL are different from the HS-LPL one because these
enzymes act in the absence of hormonal action, and so the regulatory step in the
lipolysis is that activated by HS-LPL. The TG, and definitly the contained fatty
acids represset the concentrated stores of metabolic energy in comparison with
that yeilded from carbohydrate and protein, the two other major nutrients
because fatty acids have the highly reduced hydrogen and are largly
anhydrous.
The next note is who the human body deals with the products of the
metabolized TG; the fatty acids and glycerol. The fatty acids formed in
adipocytes are either reuesd again in adipocytes(intraadipocytes) for the
synthesis of new molecules of TG(lipogenesis) as the two pathways of liogenesis
and lipolysis are in dynamic state. The other fate of fatty acids is their release of
the formed fatty acids in the circulation, where they are combined with blood
albumin to be able to transport in the blood??. The transporting of fatty acids
in the blood is restricteted to those of long chain, while of the short chain not
need for albumin carrier. Fatty acids are transported to different tissues,
predemonantly the liver and skeletal and cardiac muscles.The Red Blood Cells
and the Brain are the only organs that not uesd the fatty acids as fuel
irrespective of their blood levels because of lack of RBCs of mitochondria and
inability of fatty acids to pass the blood-brain carrier. In the liver the entered
fatty acids are either used for resynthesis of TG in the cytoplasm and
transported again to adipose tissues as VLDL:
Fatty acids+ glycerol……..» TG………….» VLDL…….» adipose tissues(stored
metabolic enery).
The second pathway of the entered saturated fatty acids is their utilization in
the production of chemical energy ATP. The latter pathway occurred in the
liver and other tissues, mainly Muscles; cardiac and skeletal. Muscles used fatty
acids even in the presence of glucose as a source of energy which spare the
glucose for other tissues dependent on glucose. The pathway of utilization of
saturated (even C2, C4,…C16,….C20….) fatty acids is the oxidation pathway
which includes; alpha(α), beta(β), and gamma(γ). The β-oxidation pathway is
the major pathway of production of energy ATP which occurred in the
mitochondria(definitly matrix of mitochondria). This means that the entered
fatty acids must be transported from the cytoplasm in the mitochondria across
its membrane, the shorter and medium chain fatty acids can pass into
mitochondria directly without need for any transporter system. The long chain
fatty acids cannot pass cross the mitochondrial memebrane MM by themselves,
but only in the presence of transporter system referred to as Carnitine system.
So, the long chain fatty acids,more than 12 C cannot be oxidized to produce
energy only in the presence of carnitine system. Carnitine system consisted of
three enzymes and carnitine substance; carnitineacyle transferase I CAT-I,
carnitine acylcarnitine translocase, and carnitine acyl transferase II ACT-II.
The first step in oxidation of saturated (Even)fatty acids is the formation of
active form Fatty acyl-CoA:
Thiokinase
Fatty acids RCCOH………………………..» Fatty acyl CoA (RCOCoA)
ATP……AMP+PPi
This reaction is activated by the Thiokinase(Fatty acyl CoAsynthetase enzyme),
an outer M M enzyme location, this reaction occurred in the cytoplasm and
then transported across the MM :
Carnitine Transporter System :
CAT-I
Fatty acylCoA (cytoplasm)…………………» Fattyacyl carnitine(interMM space)
↑
.
Carnitine molecule(inter MM space)
.
carnitine→
(from Inner Mitoch. Matrix)
.
.
BY
.
Carnitineacylacarnitine Translocase enzyme
.
.
↓
Fattyacylcarnitine(inner MM;Mitoch. Matrix)
.
.
carnitine ←
←.
To inter MM space
.
.
.
.
↓
Fatty acyl CoA(Inner MM; Mitoch. matrix)
Ready for β-oxidation pathway
The Mitoch. Matrix fatty acyl CoA is now ready for reactions of β-oxidation
pathway:
RCO CoA(e.g; CH3(CH2)12CH2-CH2COCoA Palmitoylacyl CoA
.
.
AcylCoA Dehydrogenase. NAD…»NADH H» Respiratory
.
chain system
.
↓
CH3(CH2)12CH=CHCOCoA unsaturated fatty acyl
Enoyl CoA
Hydratase
H2O→
.
.
.
↓
(Enoyl CoA)
CH3(CH2)12CHOHCH2COCoA β-hydroxyfattyacyl CoA
.
Β-hydroxyfattyacylCoA Dehydrogenase
.
FAD…»FADH2»Respiratory
.
Chain system
↓
CH3(CH2)12COCH2COCoA β-Ketoacyl CoA
CoA.SH →
.
Thiolase
.
↓→→ CH3CoA acetyl CoA…» CAC
↓
pathway
CH3(CH2)12CoA Fattyacyl CoA(less-C2)
So with each run or turn of these reaction of β-oxidation of fattyacyl, the
product is the fattyacyl of less two carbon unit C2 of the original
one(C16….»C14) Plus the Acetyl-CoA which is the substrate of Citric Acid
Cycle CAC pathway with end products are CO2+H2O. These above steps or
reactions continue with removal of C2 until the remainder fattyacyl is
CH3COCH2CoA C4 which splitted by thiolase enzyme into two C2
2Aetyl
CoA. So, for fattyacyl C16 Palmitoyl acyl CoA there is need for 7 turn of βoxidation steps or reaction:
C16…1.C14…2…C12…3….C10…4.C8…5.C6…6….C4→ 2 Acetyl CoA
↓
↓
↓
↓
↓
↓
Acetyl CoA AcetylCoA Acetyl CoA Acetyl CoA Acetyl CoA Acetyl CoA
So, the overall pathway of β-oxidation of C16 palmitic saturated acid leads to
formation of 8 aceyl CoA by 7 turn or cycles(consisted of 4 steps or reactions),
these aceyl CoA molecules then enter CAC with formation of 3NADH H, 1
FADH2, and 1 GTP(GTP….ATP). The net amounts of ATPs that yeilded from
complete oxidation of palmitic acid C16; β-oxidation+CAC is:
7 turn of β-oxidation…….» 7 NADH H……..» 7×3=21 ATP
7 FADH2 ………» 7× 2= 14 ATP
8 Acetyl CoA
CAC
8(3NADH H+ FADH2+GTP=12 ATP)» 96 ATP
The total ATP s= 131 ATPs
-2ATPs
The net amounts of ATPs: 129 ATPs
Calculate the amounts of NADH H, FADH2 and ATPs from 1. Β-oxidation 2.
Complete oxidation to CO2+H2O??.
The regulatory step in β-oxidation pathway is at the CAT-I step.Malonyl CoA,
the intermediate of fatty acids synthesis, is the inhibitor of by β-oxidation by
inhibiting of CPT-I enzyme of carnitine system. The inhibiting of CPT-I results
in preventing the fatty (long chain)acyl CoA from entry into mitochondrial
matrix and so from oxidation in β-pathway. So, in case of CHO intake(post
CHO meal, or high caloric intake) the levels of glucose and insulin are high, the
amounts of citrate and so of Malonyl CoA are also high, the synthesized
malonyl CoA will inhibits the CPT-I and therefore the newly synthesized fatty
acids cannot not pass into the mitochondrial matrix and so will be reacted with
glycerol to form TG, → VLDL which secreted into circulation and transported
to the adipose tissues(energy storage form) and muscles(energy source). The
vice versa, in prolonged fasting state the glucose and insulin are low, the
malonyl CoA low, the inhibition of CPT-I will be removed and the entered fatty
acids into the cytoplasm will be proceed into the β-oxidation pathway.
Carnitine substance are found in meat(exogenous sources). In human body
carnitine is synthesizes endogenously fro Lysine and Methionine amion acids in
the liver and kidney but not the muscles. However, the synthesized carnitine
will be transported to the muscles; cardiac and skeletal where they are used foe
fatty acids oxidation to produce the ATP. About 97 % of the body ΄s carnitine
is predominant in the muscles.
Disorders of Carnitine Deficiences and β-Oxidation Impairment:
Carnitine deficiency which may be primary and secondary leads to decrease
utilization of long chain fatty acid LCFA as source of energy, pariculary in the
muscles, with dependent mainly on the glucose and so the occurrence of
Hypoglycemia.The primary causes are:1. genetic defect or deficiency of CAT-I
which affects mainly the Liver and prevent it from utilization of LCFA to
produce of energy and so decrease its ability for formation of glucose and spare
it for those glucose dependent tissues(RBCs, brain, kidney…). The outcome of
this defect is hypoglycemia, coma and death. 2. Genetic defect or deficiency of
CAT-II which affects mainly the muscles(cardiac and skeletal) and leads to
defect in utilization of LCFA with cardiomyopathy, muscle weakness and
myoglobinemia after prolonged exercise??. Tratment, by low lipid or fat diet
and rich CHO , but high amounts of short and medium fatty acids is
recommended ??. supplementation with carnitine is also recommended in case
of carnitine deficiency.
Impairment of β-oxidation including the genetic deficiency of Medium Chain
Fatty Acyl CoA Dehydrogenaes enzyme MCAD leads to defect in β-oxidation
of MCFA which is predominant in milk and so affects mainly the infants who
are dependent on milk in their nourishment or feeding. MCAD disease is
autosomal recessive disease and is the most common inborn error of
metabolism and the most common inborn error of fatty acid oxidation. MCAD
disease is considered as one of the sudden infant death syndrome SIDS or Reye
syndrome.
The oxidation of Odd number(C1,…C3,….C15,…..) saturated fatty acids also
proceeded as the even number in β-oxidation, but the pathway terminates with
C5which splitted into the acetyl CoA + C3 Propionyl CoA. The acetyl CoA also
oxidized in CAC, but the propionyl CoA is metabolized by different pathway as
follows:
Carboxylase CO2
Propinoyl CoA………………………↓…..» D-Methyl malonyl CoA
COOATP……..ADP+Pi
.
|
Biotin Co E
.
Racemase .
.
↓
L-Methylmalonyl CoA
CoE Vit. B12 .
.
Mutase .
↓
Gluconeogenesis
← Succinyl CoA
(Formation of glucose from fatty acids) ↓
CAC
H- C-CH3
|
CO CoA
COO|
H3C -C-H
|
CO CoA
So the only saturated fatty acids that can be used in production of
glucose(gluconeogenesis pathway) is the odd nembeer fatty acids, while the
naturally endogenous synthesized fatty acids(the even number) cannot be used
as sourcer of gluconeogenesis??.
Oxidation of very long saturated fatty acids VLCFA which predominant in
nervous tissues is also by β-oxidation but in Peroxisome organelle instead of
mitochondria, this pathway terminates in shorter fatty acids and acetyl CoA
which transferred to the mitochondria for further complete oxidation by also βoxidation and subsequent CAC. Zellweger syndrome is characterized by
accumulation of VLCFAs in tissues and blood due to defect in biogenesis of
peroxisome organelle(congenital defect) or genetic defect in transporting of
VLCFA across the peroxisome membrane
resulting in X-linked
Adrenoleukodystrophy.
Phytanic acid is branched fatty acid with methyl group at C3(β position). This
fatty acid cannot be oxidized by Fatty acyl CoA DH of β-oxidaton, instead is
firstly oxidized by α-hydoxylase enzyme to produce the acyl-CoA derivatives
which then
proceeded in β-oxidation pathway. Refsum disease is characterized by
primarily the neurologic symptoms due to accumulation of phytanic acid in
blood and tissues because of genetic deficiency of α-hydroxylase enzyme and its
treatment is by dietary restriction of this fatty acid.