Intermediary Metabolism II
1- Write an account of oxidative phosphorylation.
Oxidative phosphorylation (or OXPHOS in short) is a metabolic pathway that uses energy released by
the oxidation ofnutrients to produce adenosine triphosphate (ATP). Although the many forms of life on
earth use a range of different nutrients, almost all aerobic organisms carry out oxidative phosphorylation
to produce ATP, the molecule that supplies energy tometabolism. This pathway is probably so pervasive
because it is a highly efficient way of releasing energy, compared to alternative fermentation processes
such as anaerobic glycolysis.
During oxidative phosphorylation, electrons are transferred from electron donors to electron
acceptors such as oxygen, inredox reactions. These redox reactions release energy, which is used to
form ATP. In eukaryotes, these redox reactions are carried out by a series of protein
complexes within mitochondria, whereas, in prokaryotes, these proteins are located in the cells' inner
membranes. These linked sets of proteins are called electron transport chains. In eukaryotes, five main
protein complexes are involved, whereas in prokaryotes many different enzymes are present, using a
variety of electron donors and acceptors.
The energy released by electrons flowing through this electron transport chain is used to transport
protons across the inner mitochondrial membrane, in a process called chemiosmosis. This
generates potential energy in the form of a pH gradient and an electrical potential across this membrane.
This store of energy is tapped by allowing protons to flow back across the membrane and down this
gradient, through a large enzyme called ATP synthase. This enzyme uses this energy to generate ATP
from adenosine diphosphate (ADP), in a phosphorylation reaction. This reaction is driven by the proton
flow, which forces the rotation of a part of the enzyme; the ATP synthase is a rotary mechanical motor.
Although oxidative phosphorylation is a vital part of metabolism, it produces reactive oxygen species such
as superoxide andhydrogen peroxide, which lead to propagation of free radicals, damaging cells and
contributing to disease and, possibly, aging(senescence). The enzymes carrying out this metabolic
pathway are also the target of many drugs and poisons that inhibittheir activities.
Oxidative phosphorylation works by using energy-releasing chemical reactions to drive energy-requiring
reactions: The two sets of reactions are said to be coupled. This means one cannot occur without the
other. The flow of electrons through the electron transport chain, from electron donors such
as NADH to electron acceptors such as oxygen, is an exergonic process – it releases energy, whereas
the synthesis of ATP is an endergonic process, which requires an input of energy. Both the electron
transport chain and the ATP synthase are embedded in a membrane, and energy is transferred from
electron transport chain to the ATP synthase by movements of protons across this membrane, in a
process called chemiosmosis.[1] In practice, this is like a simple electric circuit, with a current of protons
being driven from the negative N-side of the membrane to the positive P-side by the proton-pumping
enzymes of the electron transport chain. These enzymes are like a battery, as they perform work to drive
current through the circuit. The movement of protons creates an electrochemical gradient across the
membrane, which is often called the proton-motive force. It has two components: a difference in proton
concentration (a H+ gradient, ΔpH) and a difference in electric potential, with the N-side having a negative
charge.[2]
ATP synthase releases this stored energy by completing the circuit and allowing protons to flow down the
electrochemical gradient, back to the N-side of the membrane.[3] This kinetic energy drives the rotation of
part of the enzymes structure and couples this motion to the synthesis of ATP.
The two components of the proton-motive force are thermodynamically equivalent: In mitochondria, the
largest part of energy is provided by the potential; in alkaliphile bacteria the electrical energy even has to
compensate for a counteracting inverse pH difference. Inversely, chloroplasts operate mainly on ΔpH.
However, they also require a small membrane potential for the kinetics of ATP synthesis. At least in the
case of the fusobacterium P. modestum it drives the counter-rotation of subunits a and c of the F0 motor
of ATP synthase.[2]
The amount of energy released by oxidative phosphorylation is high, compared with the amount produced
by anaerobic fermentation. Glycolysis produces only 2 ATP molecules, but somewhere between 30 and
36 ATPs are produced by the oxidative phosphorylation of the 10 NADH and 2 succinate molecules made
by converting one molecule of glucose to carbon dioxide and water,[4] while each cycle of beta
oxidation of a fatty acid yields about 14 ATPs. These ATP
The electron transport chain carries both protons and electrons, passing electrons from donors to
acceptors, and transporting protons across a membrane. These processes use both soluble and proteinbound transfer molecules. In mitochondria, electrons are transferred within the intermembrane space by
the water-soluble electron transfer protein cytochrome c.[6] This carries only electrons, and these are
transferred by the reduction and oxidation of an iron atom that the protein holds within a heme group in its
structure. Cytochrome c is also found in some bacteria, where it is located within theperiplasmic space.[7]
Within the inner mitochondrial membrane, the lipid-soluble electron carrier coenzyme Q10 (Q) carries
both electrons and protons by a redox cycle.[8]This small benzoquinone molecule is very hydrophobic, so
it diffuses freely within the membrane. When Q accepts two electrons and two protons, it becomes
reduced to the ubiquinol form (QH2); when QH2 releases two electrons and two protons, it becomes
oxidized back to the ubiquinone (Q) form. As a result, if two enzymes are arranged so that Q is reduced
on one side of the membrane and QH2 oxidized on the other, ubiquinone will couple these reactions and
shuttle protons across the membrane.[9] Some bacterial electron transport chains use different quinones,
such as menaquinone, in addition to ubiquinone.[10]
Within proteins, electrons are transferred between flavin cofactors,[3][11] iron–sulfur clusters, and
cytochromes. There are several types of iron–sulfur cluster. The simplest kind found in the electron
transfer chain consists of two iron atoms joined by two atoms of inorganic sulfur; these are called [2Fe–
2S] clusters. The second kind, called [4Fe–4S], contains a cube of four iron atoms and four sulfur atoms.
Each iron atom in these clusters is coordinated by an additional amino acid, usually by the sulfur atom
of cysteine. Metal ion cofactors undergo redox reactions without binding or releasing protons, so in the
electron transport chain they serve solely to transport electrons through proteins. Electrons move quite
long distances through proteins by hopping along chains of these cofactors. [12] This occurs by quantum
tunnelling, which is rapid over distances of less than 1.4×10−9 m.[13]
[edit]Eukaryotic
electron transport chains
Further information: Electron transport chain and Chemiosmosis
Many catabolic biochemical processes, such as glycolysis, the citric acid cycle, and beta oxidation,
produce the reduced coenzyme NADH. This coenzyme contains electrons that have a high transfer
potential; in other words, they will release a large amount of energy upon oxidation. However, the cell
does not release this energy all at once, as this would be an uncontrollable reaction. Instead, the
electrons are removed from NADH and passed to oxygen through a series of enzymes that each release
a small amount of the energy. This set of enzymes, consisting of complexes I through IV, is called the
electron transport chain and is found in the inner membrane of the mitochondrion. Succinate is also
oxidized by the electron transport chain, but feeds into the pathway at a different point.
In eukaryotes, the enzymes in this electron transport system use the energy released from the oxidation
of NADH to pump protons across the inner membrane of the mitochondrion. This causes protons to build
up in the intermembrane space, and generates an electrochemical gradient across the membrane. The
energy stored in this potential is then used by ATP synthase to produce ATP. Oxidative phosphorylation
in the eukaryotic mitochondrion is the best-understood example of this process. The mitochondrion is
present in almost all eukaryotes, with the exception of anaerobic protozoa such as Trichomonas
vaginalis that instead reduce protons to hydrogen in a remnant mitochondrion called a hydrogenosome.[
2- Enumerate and briefly describe the glycogen storage diseases.
Glycogen storage disease (GSD, also glycogenosis and dextrinosis) is the result of defects in the
processing of glycogen synthesis or breakdown within muscles, liver, and other cell types.[1] GSD has two
classes of cause: genetic and acquired. Genetic GSD is caused by any inborn error of
metabolism (genetically defective enzymes) involved in these processes. In livestock, acquired GSD is
caused by intoxication with the alkaloidcastanospermine.[2]
Overall, according to a study in British Columbia, approximately 2.3 children per 100 000 births (1 in
43,000) have some form of glycogen storage disease.[3] In the United States, they are estimated to occur
in 1 per 20,000-25,000 births.[4] A Dutch study estimated it to be 1 in 40,000.[5]
[edit]Types
There are eleven (11) distinct diseases that are commonly considered to be glycogen storage diseases
(some previously thought to be distinct have been reclassified). (Although glycogen synthase deficiency
does not result in storage of extra glycogen in the liver, it is often classified with the GSDs as type 0
because it is another defect of glycogen storage and can cause similar problems.)

GSD type VIII: In the past, considered a distinct condition.[6] Now classified with VI.[7] Has been
described as X-linked recessive.[8]

GSD type X: In the past, considered a distinct condition.[9][10] Now classified with VI

Glycogen storage disease type I (GSD I) or von Gierke's disease, is the most common of
the glycogen storage diseases. This genetic disease results from deficiency of
the enzyme glucose-6-phosphatase. This deficiency impairs the ability of the liver to produce
free glucose from glycogen and fromgluconeogenesis. Since these are the two
principal metabolic mechanisms by which the liver supplies glucose to the rest of the body during
periods offasting, it causes severe hypoglycemia. Reduced glycogen breakdown results in
increased glycogen storage in liver and kidneys, causing enlargement of both. Both organs
function normally in childhood but are susceptible to a variety of problems in the adult years.
Other metabolic derangements includelactic acidosis and hyperlipidemia. Frequent or continuous
feedings of cornstarch or other carbohydrates are the principal treatment. Other therapeutic
measures may be needed for associated problems.

The disease is named after Edgar von Gierke,[1][2] the German doctor who discovered it.
3- Write an account of ketosis.
etosis (
/kɨˈtoʊsɨs/) is a state of elevated levels of ketone bodies in the body.[1] It is almost always
generalized throughout the body, withhyperketonemia, that is, an elevated level of ketone bodies in the
blood. Ketone bodies are formed by ketogenesis when the liver glycogen stores are depleted. The ketone
bodies acetoacetate and β-hydroxybutyrate are used for energy. When glycogen stores are not available
in the cells, fat (triacylglycerol) is cleaved to provide 3 fatty acid chains and 1 glycerol molecule in a
process calledlipolysis. Most of the body is able to use fatty acids as an alternative source of energy in a
process called beta-oxidation. One of the products of beta-oxidation is acetyl-CoA, which can be further
used in the Krebs cycle. During prolonged fasting or starvation, acetyl-CoA in the liver is used to
produceketone bodies instead, leading to a state of ketosis.
During starvation or a long physical training session, the body starts using fatty acids instead of glucose.
The brain cannot use long-chain fatty acids for energy because they are completely albumin-bound and
cannot cross the blood-brain barrier. Not all medium-chain fatty acids are bound to albumin. The unbound
medium-chain fatty acids are soluble in the blood and can cross the blood-brain barrier.[2] The ketone
bodies produced in the liver can also cross the blood-brain barrier. In the brain, these ketone bodies are
then incorporated into acetyl-CoA and used in the citric acid cycle.
The ketone body acetoacetate will slowly decarboxylate into acetone, a volatile compound that is both
metabolized as an energy source and lost in thebreath and urine.
[edit]Ketoacidosis
Main article: Ketoacidosis
Ketone bodies are acidic, but acid-base homeostasis in the blood is normally maintained
through bicarbonate buffering, respiratory compensation to vary the amount of CO2 in the bloodstream,
hydrogen ion absorption by tissue proteins and bone, and renal compensation through increased
excretion of dihydrogen phosphate and ammonium ions.[3] Prolonged excess of ketone bodies can
overwhelm normal compensatory mechanisms, leading to acidosis if blood pH falls below 7.35.
There are two major causes of ketoacidosis:

Most commonly, ketoacidosis is diabetic ketoacidosis (DKA), resulting from increased fat metabolism
due to a shortage of insulin. It is associated primarily with type I diabetes, and may result in adiabetic
coma if left untreated.[4]

Alcoholic ketoacidosis (AKA) presents infrequently, but can occur with acute alcohol intoxication,
most often following a binge in alcoholics with acute or chronic liver or pancreatic disorders. Alcoholic
ketoacidosis occurs more frequently following methanol or ethylene glycol intoxication than following
intoxication with uncontaminated ethanol.[5]
A mild acidosis may result from prolonged fasting or when following a ketogenic diet.[6]
[edit]Diet
If the diet is changed from a highly glycemic diet to a diet that does not provide sufficient carbohydrate to
replenish glycogen stores, the body goes through a set of stages to enter ketosis. During the initial stages
of this process, blood glucose levels are maintained through gluconeogenesis, and the adult brain does
not burn ketones. However, the brain makes immediate use of ketones for lipid synthesis in the brain.
After about 48 hours of this process, the brain starts burning ketones in order to more directly use the
energy from the fat stores that are being depended upon, and to reserve the glucose only for its absolute
needs, thus avoiding the depletion of the body's protein store in the muscles. [7]
Ketosis is deliberately induced by use of a ketogenic diet as a medical intervention in cases of
intractable epilepsy.[6] Other uses of low-carbohydrate diets remain controversial.[8][9]
[edit]Diagnosis
Whether ketosis is taking place can be checked by using special urine test strips such as Ketostix. The
strips have a small pad on the end which is dipped in a fresh specimen of urine. Within a matter of
seconds, the strip changes color indicating the level of ketone bodies detected, which reflects the degree
of ketonuria, which, in turn, can be used to give a rough estimation of the level of hyperketonemia in the
body (see table below). Normal serum reference ranges for ketone bodies are 0.5-3.0 mg/dL, equivalent
to 0.05-0.29 mmol/L.[10]
Also, when the body is in ketosis, subjects often smell of acetone
4- Describe the causes, pathogenesis and consequences of phenyl ketonuria.
Phenylketonuria (PKU) is an autosomal recessive metabolic genetic disorder characterized by a
mutation in the gene for the hepatic enzymephenylalanine hydroxylase (PAH), rendering it
nonfunctional.[1]:541 This enzyme is necessary to metabolize the amino acid phenylalanine (Phe) to the
amino acid tyrosine. When PAH activity is reduced, phenylalanine accumulates and is converted
into phenylpyruvate (also known as phenylketone), which is detected in the urine.[2]
Since its discovery, there have been many advances in its treatment. It can now be successfully
managed by the patient under ongoing medical supervision to avoid the more serious side effects. If,
however, the condition is left untreated, it can cause problems with brain development, leading to
progressivemental retardation, brain damage, and seizures. Early cases of PKU were treated with a lowphenylalanine diet. More recent research has now shown that diet alone may not be enough to prevent
the negative effects of elevated phenylalanine levels. Optimal treatment involves maintaining blood Phe
levels in a safe range while monitoring diet and cognitive development. There is no cure for PKU, but
patients who are diagnosed early and maintain a strict diet can have a normal life span with normal
mental development.
Phenylketonuria was discovered by the Norwegian physician Ivar Asbjørn Følling in 1934[3] when he
noticed hyperphenylalaninemia (HPA) was associated with mental retardation. In Norway, this disorder is
known as Følling's disease, named after its discoverer.[4] Dr. Følling was one of the first physicians to
apply detailed chemical analysis to the study of disease. His careful analysis of the urine of two affected
siblings led him to request many physicians near Oslo to test the urine of other affected patients. This led
to the discovery of the same substance he had found in eight other patients. He conducted tests and
found reactions that gave rise to benzaldehyde and benzoic acid, which led him to conclude that the
compound contained a benzene ring. Further testing showed themelting point to be the same
as phenylpyruvic acid, which indicated that the substance was in the urine. His careful science inspired
many to pursue similar meticulous and painstaking research with other disorders. utations can also cause
PKU. This is an example of non-allelic genetic heterogeneity. The PAH gene is located on chromosome
12 in the bands 12q22-q24.1. More than 400 disease-causing mutations have been found in the PAH
gene. PAH deficiency causes a spectrum of disorders, including classic phenylketonuria (PKU) and
hyperphenylalaninemia (a less severe accumulation of phenylalanine). [7]
PKU is known to be an autosomal recessive genetic disorder. This means both parents must have at
least one mutated allele of the PAH gene. The child must inherit both mutated alleles, one from each
parent. Therefore, it is not impossible for a parent with the disease to have a child without it if the other
parent possesses one functional allele of the gene for PAH. Yet, a child from two parents with PKU will
inherit two mutated alleles every time, and therefore the disease.
Phenylketonuria can exist in mice, which have been extensively used in experiments into an effective
treatment for it.[8] The macaque monkey's genome was recently sequenced, and the gene encoding
phenylalanine hydroxylase was found to have the same sequence that, in humans, would be considered
the PKU mutation.[9]
[edit]Tetrahydrobiopterin-deficient hyperphenylalaninemia
A rarer form of hyperphenylalaninemia occurs when PAH is normal, but there is a defect in the
biosynthesis or recycling of the cofactor tetrahydrobiopterin (BH4) by the patient.[10] This cofactor is
necessary for proper activity of the enzyme. The coenzyme (called biopterin) can be supplemented as
treatment. Those who suffer from PKU must be supplemented with Tyrosine to account for Phenylalanine
Hydroxylase deficiency in converting Phenylalanine to Tyrosine sufficiently. Dihydrobiopterin Reductase
activity is to replenish Dihydrobiopterin back into its Tetrahydrobiopterin form, which is an important
cofactor in many metabolic reactions in amino acid metabolism. Those with this deficiency may produce
sufficient levels of Phenylalanine Hydroxylase, but since Tetrabiopterin is a cofactor for Phenylalanine
Hydroxylase activity, deficient Dihydrobiopterin Reductase renders any Phenylalanine Hydroxylase
enzyme produced unable to use Phenylalanine to produce Tyorosine. Tetrahydrobiopterin is also a
cofactor in the production of L-DOPA from Tyrosine and 5-Hydroxy-L-Tryptophan from Tryptophan, which
must also be supplemented as treatment in addition to the supplements for Classical PKU.
Levels of dopamine can be used to distinguish between these two types. Tetrahydrobiopterin is required
to convert phenylalanine to tyrosine, but it is also required to convert tyrosine to L-DOPA (via the
enzyme tyrosine hydroxylase), which in turn is converted to dopamine. Low levels of dopamine lead to
high levels of prolactin. By contrast, in classical PKU, prolactin levels would be relatively
normal.Tetrahydrobiopterin deficiency can be caused by defects in four different genes. These types are
known as HPABH4A, HPABH4B, HPABH4C, and HPABH4D.[11]
[edit]Metabolic
pathways
The enzyme phenylalanine hydroxylase normally converts the amino acid phenylalanine into the amino
acid tyrosine. If this reaction does not take place, phenylalanine accumulates and tyrosine is deficient.
Excessive phenylalanine can be metabolized into phenylketones through the minor route,
a transaminase pathway with glutamate. Metabolites
include phenylacetate, phenylpyruvate andphenethylamine.[12] Elevated levels of phenylalanine in the
blood and detection of phenylketones in the urine is diagnostic, however most patients are diagnosed via
newborn screening.
Phenylalanine is a large, neutral amino acid (LNAA). LNAAs compete for transport across the blood-brain
barrier (BBB) via the large neutral amino acid transporter (LNAAT). If phenylalanine is in excess in the
blood, it will saturate the transporter. Excessive levels of phenylalanine tend to decrease the levels of
other LNAAs in the brain. However, as these amino acids are necessary for protein and neurotransmitter
synthesis, Phe buildup hinders the development of the brain, causing mental retardation.[13]
[edit]Treatment
If PKU is diagnosed early enough, an affected newborn can grow up with normal brain development, but
only by managing and controlling Phe levels through diet, or a combination of diet and
medication. Optimal health ranges (or "target ranges") are between 120 and 360 µmol/L, and aimed to be
achieved during at least the first 10 years.[14] When Phe cannot be metabolized by the body, abnormally
high levels accumulate in the blood and are toxic to the brain. When left untreated, complications of PKU
include severe mental retardation, brain function abnormalities, microcephaly, mood disorders, irregular
motor functioning, and behavioral problems such as attention deficit hyperactivity disorder.
All PKU patients must adhere to a special diet low in Phe for optimal brain development. "Diet for life" has
become the standard recommended by most experts. The diet requires severely restricting or eliminating
foods high in Phe, such as meat, chicken, fish, eggs, nuts, cheese, legumes, milk and other dairy
products. Starchy foods, such as potatoes, bread, pasta, and corn, must be monitored. Infants may still
be breastfed to provide all of the benefits of breastmilk, but the quantity must also be monitored and
supplementation for missing nutrients will be required. The sweetener aspartame, present in many diet
foods and soft drinks, must also be avoided, as aspartame consists of two amino acids: phenylalanine
and aspartic acid.
Supplementary infant formulas are used in these patients to provide the amino acids and other necessary
nutrients that would otherwise be lacking in a low-phenylalanine diet. As the child grows up these can be
replaced with pills, formulas, and specially formulated foods. (Since Phe is necessary for the synthesis of
many proteins, it is required for appropriate growth, but levels must be strictly controlled in PKU patients.)
In addition, tyrosine, which is normally derived from phenylalanine, must be supplemented.
The oral administration of tetrahydrobiopterin (or BH4) (a cofactor for the oxidation of phenylalanine) can
reduce blood levels of this amino acid in certain patients.[15][16] The company BioMarin
Pharmaceutical has produced a tablet preparation of the compound sapropterin dihydrochloride (Kuvan),
which is a form of tetrahydrobiopterin. Kuvan is the first drug that can help BH4-responsive PKU patients
(defined among clinicians as about 1/2 of the PKU population) lower Phe levels to recommended
ranges.[17] Working closely with a dietitian, some PKU patients who respond to Kuvan may also be able to
increase the amount of natural protein they can eat.[18] After extensive clinical trials, Kuvan has been
approved by the FDA for use in PKU therapy. Some researchers and clinicians working with PKU are
finding Kuvan a safe and effective addition to dietary treatment and beneficial to patients with PKU. [
5- Write an account on the dark reactions of photosynthesis.
Photosynthesis in plants takes place
in chloroplasts. Photosynthesis includes lightdependent reactions and reactions that are not
directly energized by light.
The structure of a chloroplast is shown on p. 872
of Biochemistry, by Voet & Voet, 3rd Edition, and
schematically represented at right.
In the photosynthetic light reactions, energy of light is conserved as as "high energy"
phosphoanhydride bonds of ATP, and as reducing power of NADPH. The proteins
and pigments responsible for the photosynthetic light reaction are associated with
the thylakoid (grana disk) membranes. The light reaction pathways will not be
presented here.
The Calvin Cycle, earlier designated the photosynthetic "dark reactions" pathway, is
now referred to as the carbon reactions pathway. In this pathway, the free energy of
cleavage of ~P bonds of ATP, and reducing power of NADPH, are used to fix and
reduce CO2 to form carbohydrate. Enzymes and intermediates of the Calvin Cycle are
located in the chloroplast stroma, a compartment somewhat analogous to
the mitochondrial matrix.
Ribulose Bisphosphate Carboxylase (RuBP Carboxylase) catalyzes CO2 fixation:
ribulose-1,5-bisphosphate + CO2  2 copies of 3-phosphoglycerate
Because it can alternatively catalyze an oxygenase reaction (discussed below), the
enzyme is also called RuBP Carboxylase/Oxygenase (RuBisCO). It is the most
abundant enzyme on earth. The RuBP Carboxylasereaction mechanism is presented
on p. 900.
Extraction of a proton
from C3 of ribulose1,5-bisphosphate
(RuBP, below left)
promotes formation of
an endiolate
intermediate.
Nucleophilic attack on
CO2 is proposed to
yield a -keto acid
intermediate, that
reacts with water and
cleaves to form 2
molecules of 3phosphoglycerate.
Transition state analogs of the postulated -keto acid intermediate
bind tightly to the enzyme and inhibit its activity. Examples include 2carboxyarabinitol-1,5-bisphosphate (CABP, at right)
and carboxyarabinitol-1-phosphate (CA1P).
RuBP Carboxylase in plants is a complex (L8S8) of:


8 large catalytic subunits (L, 477 amino acid residues)
8 small subunits (S, 123 amino acid residues).
Some bacteria contain only the large subunit, with the smallest functional unit being a
homodimer, L2. Roles of the small subunits have not been clearly defined, although
there is some evidence that interactions between large and small subunits may
regulate catalysis.
At right are 2 views of
spinach RuBisCO (RuBP
Carboxylase), with large
subunits colored blue or
cyan, and small subunits
colored red.
Large subunits within RuBisCO are arranged as antiparallel
dimers, with the N-terminal domain of one monomer
adjacent to the C-terminal domain of the other monomer.
Each active site is at an interface between monomers within
an L2 dimer, explaining the minimal requirement for a
dimeric structure.
The substrate binding site is at the mouth of an -barrel
domain of the large subunit. Most active site residues are
polar, including some charged amino acids (e.g., Thr, Asn,
Glu, Lys).
"Active" RuBP Carboxylase includes
a carbamate group, that binds an
essential Mg++ at the active site.
The carbamate forms by reaction of
HCO3 with the -amino group of
a lysine residue of RuBP
Carboxylase, in the presence of
Mg++. HCO3 that reacts to form the
carbamate group is distinct from
CO2 that binds to RuBP Carboxylase
as substrate.
The active site Mg++ bridges between
oxygen atoms of the carbamate and
the substrate CO2.
Binding of either the normal substrate ribulose-1,5-bisphosphate or a transition state
analog to RuBP Carboxylase causes a conformational change to a "closed"
conformation in which access of solvent water to the active site is blocked.
RuBP Carboxylase (RuBisCO) can spontaneously deactivate by decarbamylation. In
the absence of the carbamate group, RuBisCO tightly binds ribulose bisphosphate
(RuBP) or another sugar phosphate at the active site as a "dead end" complex, with
the closed conformation, and is inactive in catalysis. In order for the carbamate to
reform, the enzyme must undergo transition to the open conformation.
RuBP Carboxylase Activase, an ATP hydrolyzing (ATPase) enzyme, causes a
conformational change in RuBP Carboxylase from closed to open form. This allows
release of tightly bound RuBP or other sugar phosphate from the active site, and
carbamate formation. Since photosynthetic light reactions produce ATP, the ATP
dependence of RuBisCO activation provides a mechanism for light-dependent
activation of the enzyme.
RuBP Carboxylase Activase is a member of the AAA family of ATPases, many of
which have chaperone-like roles. The activase is a large multimeric protein complex
that may surround RuBP Carboxylase while inducing the conformational change to
the open state.
Explore at right the bound carbamate and reaction products at the active site of RuBisCO.
6- Explain in detail glycol sis. Add a note on the energetic of glycol sis.
Glycolysis (from glycose, an older term [1] for glucose + -lysis degradation) is the metabolic pathway that
converts glucose C6H12O6, into pyruvate, CH3COCOO− + H+. The free energyreleased in this process is
used to form the high-energy compounds ATP (adenosine triphosphate) and NADH (reduced
nicotinamide adenine dinucleotide).
Glycolysis is a definite sequence of ten reactions involving ten intermediate compounds (one of the steps
involves two intermediates). The intermediates provide entry points to glycolysis. For example, most
monosaccharides, such as fructose, glucose, andgalactose, can be converted to one of these
intermediates. The intermediates may also be directly useful. For example, the intermediate
dihydroxyacetone phosphate (DHAP) is a source of the glycerol that combines with fatty acids to form fat.
It occurs, with variations, in nearly all organisms, both aerobic and anaerobic. The wide occurrence of
glycolysis indicates that it is one of the most ancient known metabolic pathways.[2] It occurs in
the cytosol of the cell.
The most common type of glycolysis is the Embden-Meyerhof-Parnas pathway (EMP pathway), which
was first discovered by Gustav Embden, Otto Meyerhof and Jakub Karol Parnas. Glycolysis also refers to
other pathways, such as the Entner–Doudoroff pathwayand various heterofermentative and
homofermentative pathways. However, the discussion here will be limited to the Embden-Meyerhof
pathway.
The entire glycolysis pathway can be separated into two phases [3]:
1. The Preparatory Phase - in which ATP is consumed and is hence also known as the investment
phase
2. The Pay Off Phase - in which ATP is produced.
The use of symbols in this equation makes it appear unbalanced with respect to oxygen atoms, hydrogen
atoms, and charges. Atom balance is maintained by the two phosphate (Pi) groups:[4]

each exists in the form of a hydrogen phosphate anion (HPO42-), dissociating to contribute 2
H+ overall

each liberates an oxygen atom when it binds to an ADP (adenosine diphosphate) molecule,
contributing 2 O overall
Charges are balanced by the difference between ADP and ATP. In the cellular environment, all three
hydroxy groups of ADP dissociate into -O- and H+, giving ADP3-, and this ion tends to exist in an ionic
bond with Mg2+, giving ADPMg-. ATP behaves identically except that it has four hydroxy groups, giving
ATPMg2-. When these differences along with the true charges on the two phosphate groups are
considered together, the net charges of -4 on each side are balanced.
For simple anaerobic fermentations, the metabolism of one molecule of glucose to two molecules of
pyruvate has a net yield of two molecules of ATP. Most cells will then carry out further reactions to 'repay'
the used NAD+ and produce a final product of ethanol orlactic acid. Many bacteria use inorganic
compounds as hydrogen acceptors to regenerate the NAD+.
Cells performing aerobic respiration synthesize much more ATP, but not as part of glycolysis. These
further aerobic reactions use pyruvate and NADH + H+ from glycolysis. Eukaryotic aerobic respiration
produces approximately 34 additional molecules of ATP for each glucose molecule, however most of
these are produced by a vastly different mechanism to the substrate-level phosphorylation in glycolysis.
The lower-energy production, per glucose, of anaerobic respiration relative to aerobic respiration, results
in greater flux through the pathway under hypoxic (low-oxygen) conditions, unless alternative sources of
anaerobically-oxidizable substrates, such as fatty acids, are found.
[edit]Elucidation
of the pathway
In 1860, Louis Pasteur discovered that microorganisms are responsible for fermentation. In 1897, Eduard
Buchner found that extracts of certain cells can cause fermentation. In 1905,Arthur Harden and William
Youngalong with Nick Sheppard determined that a heat-sensitive high-molecular-weight subcellular
fraction (the enzymes) and a heat-insensitive low-molecular-weight cytoplasm fraction (ADP, ATP and
NAD+ and other cofactors) are required together for fermentation to proceed. The details of the pathway
were eventually determined by 1940, with a major input from Otto Meyerhof and some years later by Luis
Leloir. The biggest difficulties in determining the intricacies of the pathway were due to the very short
lifetime and low steady-state concentrations of the intermediates of the fast glycolytic reactions.
7- Describe the ß -oxidation of fatty acids. Discuss the number of ATP's formed on
complete oxidation of one molecule of palmitic acid.
8- Write an essay on nitrogen fixation.