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Q.1- 20 Multiple choice questions

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Q.2- a) Explain the effect of substrate concentration in an enzyme catalyzed reaction referring the Michaelis- Menten’s equation, explain why the value of Km is of  interest. Sketch a graphical method for determining Km.            

Answer- For a typical enzyme, as substrate concentration is increased; Vi increases until it reaches a maximum value Vmax (Figure). When further increases in substrate concentration do not further increase Vmax the enzyme is said to be “saturated” with substrate. If a curve is plotted, the shape of the curve that relates activity to substrate concentration (Figure) is hyperbolic.

 Effect of substrate conc.

Figure- Effect of substrate concentration on reaction rate

At any given instant, only substrate molecules that are combined with the enzyme as an ES complex can be transformed into product. At points A or B, increasing or decreasing [S] therefore will increase or decrease the number of ES complexes with a corresponding change in Vi. This is called first order reaction, where the rate of the reaction is proportional to the substrate concentration.

At point C (Figure), essentially all the enzyme is present as the ES complex. Since no free enzyme remains available for forming ES, further increases in [S] cannot increase the rate of the reaction. This is called Zero order reaction, the reaction rate is independent of substrate concentration.

The Michaelis-Menten equation is a quantitative description of the relationship among the rate of an enzyme- catalyzed reaction [V1], the concentration of substrate [S] and two constants, Vmax and km (which are set by the particular equation). The symbols used in the Michaelis-Menten equation refer to the reaction rate [V1], maximum reaction rate (V max), substrate concentration [S] and the Michaelis-Menten constant (km).

 MM equation

The Michaelis-Menten equation can be used to demonstrate that at the substrate concentration that produces exactly half of the maximum reaction rate, i.e. ½ V max the substrate concentration is numerically equal to Km.

The dependence of initial reaction velocity on [S] and Km may be illustrated by evaluating the Michaelis-Menten equation under three conditions.

(1) When [S] is much less than km, the term km + [S] is essentially equal to km. Replacing Km + [S] with Km reduces the equation to


Where means approximately equal to

 Since V max and km are both constants, their ratio is a constant (k). In other words, when [S] is considerably below km, V max is proportionate to k[S]. The initial reaction velocity therefore is directly proportionate to [S].

(2) When [S] is much greater than km, the term km + [S] is essentially equal to [S]. Replacing km + [S] with [S] reduces the equation to


Thus, when [S] greatly exceeds km, the reaction velocity is maximal (V max) and unaffected by further increases in substrate concentration.

(3) When [S] = km


Equation states that when [S] equals km, the initial velocity is half-maximal.

Thus, the Michaelis constant, km is the substrate concentration at which V1 is half the maximal velocity (V max /2) attainable at a particular concentration of enzyme. Km thus has the dimensions of substrate concentration (Figure-2)

 Derivation of km

Figure-2- Plot of substrate concentration versus reaction velocity showing the value of km

Significance of Km

1) It is specific for a given enzyme for its specific substrate

2) It helps in determining the true substrate of the enzyme

3) Its value denotes the affinity of an enzyme for its substrate. Lower the km value more is the affinity of the enzyme for its substrate and vice versa.


b) Explain the mechanism of action of enzymes giving suitable examples.         

Answer- Enzymes are catalysts and increase the speed of a chemical reaction without themselves undergoing any permanent chemical change. They are neither used up in the reaction nor do they appear as reaction products.

Enzymes employ multiple mechanisms to facilitate catalysis.  The mechanism of action of enzymes can be explained by two perspectives-

1) Thermodynamic changes

2) Processes at the active site

1) Thermodynamic changes – All enzymes accelerate reaction rates by providing transition states with a lowered GF for formation of the transition states. However, they may differ in the way this is achieved.

A chemical reaction of substrate S to form product P goes through a transition state S‡ that has a higher free energy than does either S or P. (The double dagger denotes a thermodynamic property of the transition state).

The difference in free energy between the transition state and the substrate is called the Gibbs free energy of activation or simply the activation energy, symbolized by ∆G.

Note that the energy of activation, or ∆G, does not enter into the final ∆G calculation for the reaction, because the energy input required to reach the transition state is returned when the transition state forms the product. The activation energy barrier suggests how enzymes enhance reaction rate without altering ∆G of the reaction: enzymes function to lower the activation energy, or, in other words, enzymes facilitate the formation of the transition state. The combination of substrate and enzyme creates a new reaction pathway whose transition-state energy is lower than hat of the reaction in the absence of enzyme (Figure 3). The lower activation energy means that more molecules have the required energy to reach the transition state. Decreasing the activation barrier is analogous to lowering the height of a high-jump bar; more athletes will be able to clear the bar. The essence of catalysis is specific binding of the transition state.

 Activational barrier

Figure-3 Enzymes Decrease the Activation Energy. Enzymes accelerate reactions by decreasing ∆G, the free energy of activation.

2) Processes at the active site-Enzymes use various combinations of four general mechanisms to achieve dramatic catalytic enhancement of the rates of chemical reactions. These are as follows-

a) Catalysis by Bond Strain: In this form of catalysis, the induced structural rearrangements that take place with the binding of substrate and enzyme ultimately produce strained substrate bonds, which more easily attain the transition state. Enzymes that catalyze lytic reactions that involve breaking a covalent bond typically bind their substrates in a conformation slightly unfavorable for the bond that will undergo cleavage. The resulting strain stretches or distorts the targeted bond, weakening it and making it more vulnerable to cleavage.

b) Catalysis by Proximity and Orientation: For molecules to react, they must come within bond-forming distance of one another. The higher their concentration, the more frequently they will encounter one another and the greater will be the rate of their reaction. When an enzyme binds substrate molecules at its active site, it creates a region of high local substrate concentration. Enzyme-substrate interactions orient reactive groups and bring them into proximity with one another.

c) Catalysis Involving Proton Donors (Acids) and Acceptors (Bases): Other mechanisms also contribute significantly to the completion of catalytic events initiated by a strain mechanism, for example, the use of glutamate as a general acid catalyst (proton donor). The ionizable functional groups of aminoacyl side chains and (where present) of prosthetic groups contribute to catalysis by acting as acids or bases.

d) Covalent Catalysis: In catalysis that takes place by covalent mechanisms, the substrate is oriented to active sites on the enzymes in such a way that a covalent intermediate forms between the enzyme or coenzyme and the substrate. One of the best-known examples of this mechanism is that involving proteolysis by serine proteases, which include both digestive enzymes (trypsin, chymotrypsin, and elastase) and several enzymes of the blood clotting cascade. These proteases contain an active site serine whose R group hydroxyl forms a covalent bond with a carbonyl carbon of a peptide bond, thereby causing hydrolysis of the peptide bond. Covalent catalysis introduces a new reaction pathway whose activation energy is lower—and therefore is faster—than the reaction pathway in homogeneous solution. The chemical modification of the enzyme is, however, transient. On completion of the reaction, the enzyme returns to its original unmodified state. Its role thus remains catalytic. Covalent catalysis is particularly common among enzymes that catalyze group transfer reactions.


Q.3-a) Differentiate between simple and conjugated proteins, illustrate your answer with suitable examples.

Answer-  Simple proteins contain only amino acids as the structural components, whereas the conjugated proteins contain a non protein component knows as prosthetic group, along with constituent amino acids. The protein part in a conjugated protein is called “apo protein” and together with “prosthetic group” it is called a “holo protein”.

Examples of simple proteins are, Albumin, globulins, protamines, prolamines, glutelins, histones and scleroproteins.     

Examples of conjugated proteins are-

1) Lipoproteins – Chylomicrons, VLDL, LDL and HDL are lipoproteins, containing lipid as the prosthetic group.      

2) Glycoproteins- contain carbohydrate as the prosthetic group. When the concentration of carbohydrate is more, they are called as Mucoproteins.               

3) Phosphoproteins- Contain phosphoric acid as the prosthetic group. Casein, Ovovitellin are the examples.

4) Metallo proteins- contain metal as the prosthetic group. Alcohol dehydrogenase (contains Zn). Xanthine oxidase (Mo), Super oxide dismutase (Cu) and Aconitase (Fe) etc., mostly these metalloproteins are enzymes.             

5) Haemo proteins- contain haem as the prosthetic group. Examples include- Haemoglobin, myoglobin, Catalase, peroxidase and cytochromes etc.

6) Nucleo proteins- are DNA or RNA binding proteins. Negatively charged nucleic acids bind with Histones or other positively charged proteins to form nucleoprotein complexes.

7) Chromo proteins- Are pigmentary proteins, e.g. Visual purple, flavoproteins etc.


b) What are the salient features of an α-helix? Name the amino acids that destabilise this structure.

  • The polypeptide backbone of a helix is twisted by an equal amount about each α-carbon.
  • A complete turn of the helix contains an average of 3.6 aminoacyl residues, and the distance it rises per turn (its pitch) is 0.54 nm.
  • The R groups of each aminoacyl residue in a helix face outward (Figure).
  • Proteins contain only L-amino acids, for which a right-handed helix is by far the more stable, and
  • Only right-handed helices are present in proteins.
  • The stability of a helix arises primarily from hydrogen bonds formed between the oxygen of the peptide bond carbonyl and the hydrogen atom of the peptide bond nitrogen of the fourth residue down the polypeptide chain (Figure).
  • The ability to form the maximum number of hydrogen bonds, supplemented by van der Waals interactions in the core of this tightly packed structure, provides the thermodynamic driving force for the formation of a helix.


Amino acids that disrupt the alpha helical structure

  • Proline- Since the peptide bond nitrogen of proline lacks a hydrogen atom to contribute to a hydrogen bond; proline can only be stably accommodated within the first turn of a helix. When present elsewhere, proline disrupts the conformation of the helix, producing a bend.
  • Glycine-Because of its small size, glycine also often induces bends in helices.
  • Tryptophan with its bulky side chain also disrupts the helical structure
  • The branched amino acids like valine, Leucine and isoleucine can also not be accommodated in this configuration
  • The charged amino acids like aspartic and glutamic acid (negatively charged) and histidine, lysine and arginine (positively charged), also disrupt the helical structure by producing electrostatic interactions.

 Alpha helix

Figure- showing the hydrogen bonding to stabilize the alpha helical structure.


c) What is protein denaturation? Describe the causes and consequences of denaturation.

Denaturation is protein unfolding, disruption of the secondary, tertiary and quaternary (if present) structures, the primary structure remains intact.

Many physical (heat, mechanical mixing or vigorous shaking, repeated thawing), or chemical agents (strong acids, alkalies and salts) can bring about denaturation.

Denaturation can be reversible or irreversible (mostly it is irreversible) and a denatured protein loses its functional capacity, becomes insoluble, precipitates out and fails to migrate in the electric field.


Q.4-a) What are sugar alcohols? Give examples of such alcohols and state their biological importance.

Sugar alcohols are produced by the reduction of the carbonyl group (Aldehyde/ ketone group) of monosaccharide.

Details of Reaction-

Under specific conditions of temperature and pressure, sugars can be reduced in the presence of hydrogen. The resultant product is a polyol or sugar alcohol (alditol) but reduction of ketose sugar produces a new asymmetric carbon atom (See figure), thus two types of sugar alcohols can be produced.

 Sugar alcohol

a) Reduction of aldoses takes place at C-1 to form sugar alcohol

 Reduction of a ketose

b) Reduction of Ketose sugars takes place at C-2 to form Sugar Alcohol

Examples and Significance of some Sugar alcohols-

1)  Sorbitol-In Diabetes Mellitus excess of Glucose is converted to Sorbitol. The osmotic effect of Sorbitol is responsible for many of the complications of diabetes mellitus e.g. Cataract formation in lens. Clinically sorbitol is dehydrated and nitrated to form Isosorbide mono and dinitrate, both of which are used in Angina.

2)  Mannitol- Mannitol is also osmotically active and is used as an infusion to lower the intracranial tension by producing forced diuresis.

3)  Dulcitol- excess of galactose in galactosemia is converted to Dulcitol. The osmotic effect of Dulcitol is similar to Sorbitol and is responsible for premature cataract formation in affected patients of galactosemia.

4)  Xylitol- is produced in Uronic acid pathway of Glucose utilization; it is subsequently oxidized to produce D- Xylulose.

5) Glycerol- is produced from Glyceraldehyde. Glycerol is used for the formation of Triglycerides and phospholipids. Clinically glycerol is nitrated to form Nitro-glycerine which is used for the treatment of angina.

6) Myo- Inositol- It is hexahydroxy alcohol, also considered a vitamin It is present in the plasma membrane and acts as a second messenger for the action of hormones.

7) Ribitol- is used in the formation of vitamin B2- (Riboflavin).


b) What are homopolysaccharides? Enlist the homopolysaccharides of biological significance. Write in detail about a homopolysaccharide that can not be digested by human beings.            

Answer- Homopolysaccharides are the polymers of one type of monosaccharides that may be glucose, galactose, and fructose or N-Acetyl glucosamine.

Based on the components-

Homopolysaccharides can be-

Glucosans- polymers of glucose- Starch, glycogen, cellulose, dextrins etc

Fructosan- polymer of fructose-Inulin

Galactosan- polymer of galactose-Agar

Based on the functional significance

Storage polysaccharides- Starch, glycogen, inulin, agar and dextrins

Structural polysaccharides- Cellulose and Chitin

The homopolysaccharide that can not be digested by human beings is cellulose.

Cellulose-Cellulose is the chief constituent of plant cell walls. It is the most abundant of all carbohydrates .It is insoluble in water, gives no color with iodine and consists of β -D-glucopyranose units linked by β 1 →4 bonds (figure) to form long, straight chains strengthened by cross-linking hydrogen bonds. Mammals lack any enzyme that hydrolyzes the β 1→ 4 bonds, and so cannot digest cellulose. It is an important source of “bulk” in the diet, and the major component of dietary fiber. Microorganisms in the gut of ruminants and other herbivores can hydrolyze the linkage and ferment the products to short-chain fatty acids as a major energy source. There is some bacterial metabolism of cellulose in the human colon.            

Structure of cellulose

Figure- Glucose residues in cellulose are linked together by β 1 →4 glycosidic linkages.


c) Highlight the major differences between table, milk and malt sugar.   


Features Table sugar Milk Sugar Malt sugar
Chemical name Sucrose Lactose Maltose
Chemical components Glucose and Fructose Glucose and Galactose 2 glucose residues
Glycosidic Linkage α(1-2 ) β(1-4) α (1-4)
Reducing property(Reaction with Benedict’s reagent) Non reducing Reducing Reducing
Mutarotation can exist in α or β anomeric forms can exist in α or β anomeric forms
Reaction with phenyl hydrazine(Osazone ) No reaction Powder puff crystals Sun flower petal shaped crystals
Cleaved by Sucrase Lactase Maltase
 Biological Significance used as table sugar(sweetener) Lactose is the only carbohydrate of milk .It is synthesized by mammary glands during lactation and is the best food for infants used as a nutrient (malt extract; Hordeum vulgare); as a sweetener and as a fermentative reagent
Clinical Significance Sucrase deficiency leads to sucrose intolerance(rare) Lactase deficiency leads to lactose intolerance(very common) No disease reported


Q.5-a) Describe the chemical nature and functions of phospholipids     

a) Phospholipids-

  • Contain in addition to fatty acids and glycerol/or other alcohol, a phosphoric acid residue, nitrogen containing base and other substituents.
  • Phospholipids may be regarded as derivatives of phosphatidic acid, in which the phosphate is esterified with the —OH of a suitable alcohol.
  • They are amphipathic molecules containing a polar head and a hydrophobic portion

Classification of phospholipids

Based on nature of alcohol-

1) Glycerophospholipids- Glycerol is the alcohol group.


  • Phosphatidyl choline
  • Phosphatidyl ethanolamine
  • Phosphatidyl  serine
  • Phosphatidyl Inositol
  • Phosphatidic acid
  • Cardiolipin
  • Plasmalogen
  • Platelet activating factor
  • Phosphatidyl Glycerol

2) Sphingophospholipids- Sphingol is the alcohol group

Example- Sphingomyelin

Functions of Phospholipids

  • Components of cell membrane, mitochondrial membrane and lipoproteins
  • Participate in lipid absorption and transportation from intestine
  • Play important role in blood coagulation
  •  Required for enzyme action- especially in mitochondrial electron transport chain
  • Choline acts as a lipotropic agent
  • Membrane phospholipids acts as source of Arachidonic acid
  • Act as reservoir of second messenger- Phosphatidyl Inositol
  • Act as cofactor for the activity of Lipoprotein lipase
  • Phospholipids of myelin sheath provide insulation around the nerve fibers
  • Dipalmitoyl lecithin acts as a surfactant


b) What are essential Fatty Acids? Describe the role of such fatty acids in the maintenance of health.         

  • Polyunsaturated fatty acids such as Linoleic and Linolenic acids are essential for normal life functions. They are therefore characterized as essential fatty acids.
  • Arachidonic acid is considered as semi essential fatty acid since it can be synthesized from Linoleic acid.
  • Essential polyunsaturated fatty acids can be classified as belonging to one of two “families”, the omega-6 family or the omega-3 family.

Significance of essential fatty acids

  • Components of cell membranes, structural elements of gonads and mitochondrial membrane
  • Required for brain growth and development
  • Precursors of Eicosanoids
  • Play important role in vision
  • They have a cardio protective role- Lower serum cholesterol and increase HDL levels
  • Prevent fatty liver formation
  • Deficiencies of essential polyunsaturated fatty acids may cause a wide variety of symptoms, including retarded growth in children, reduced fertility and pathologic changes in the skin.


c) Give an example of each of the following-                                                                    

 i) Saturated fatty acid with 18 carbon atoms – Stearic acid

 ii) Glycolipid acting as a receptor for cholera toxin- GM-1

 iii) Amphipathic molecule used for delivery of drugs- Liposome

 iv)  A lipid that acts as a precursor of thromboxane- Arachidonic acid


d) Enlist the important functions of prostaglandins, and mention the causes for their limited use in clinical practice.                   

Functions of Prostaglandins

They have various roles in inflammation, fever, regulation of blood pressure, blood clotting, immune system modulation, control of reproductive processes, tissue growth, and regulation of the sleep/wake cycle.

Due to their non specific actions, PGs are not used as conventional drugs. Diarrhoea, abdominal cramps and bronchoconstriction are the common side effects of prostaglandins.

Q.6- Describe the following diseases explaining the biochemical defect, clinical manifestations, laboratory diagnosis and treatment-

 a) Gaucher disease

This disease is a multisystem lipidosis characterized by hematological changes, organomegaly and skeletal involvement, the latter is usually manifested in the form of bone pains and multiple fractures. It is the most common genetic disorder among Ashkenazi Jews. It is the commonest Lysosomal storage disease.


It is Autosomal recessive in nature.

Biochemical defect

Gaucher’s disease results from deficient activity of Lysosomal Hydrolase, β- Glucocerebrosidase. The enzyme defect results in accumulation of undegraded glycolipid in the form of Glucosyl ceramide in the cells of reticuloendothelial system. This progressive accumulation results in infiltration of bone marrow, hepatosplenomegaly and skeletal complications.

Clinical features

There are three clinical subtypes depending upon the presence of, absence of or progression of neurological complications-

1) Type-1– It accounts for 99 % of cases. The age of onset is variable, from early childhood to late adulthood. The patients present with easy bruising due to thrombocytopenia, chronic fatigue due to anemia, hepatomegaly with or without impaired liver functions. Progressive enlargement of spleen is there which can become massive. Clinical bone involvement is apparent which is manifested in the form of bone pains, or pathological fracturesThe hall mark of Gaucher’s disease is gaucher cells in the reticuloendothelial system, particularly in the bone marrow. These cells have a typical appearance, they are 20-100μm in diameter, wrinkled looking due to the presence of intracytoplasmic inclusion bodies. The presence of these cells is highly diagnostic of Gaucher’s disease.

2) Type 2- is less common, it is characterized by neurodegeneration, extreme visceral involvement and death within 2 years of life. The death is due to respiratory compromise.

3)  Type 3– is intermediate in presentation to type 1 and 2. Neurological involvement is there but occurs later in life with decreased severity as compared to Type 2.

Laboratory Diagnosis

The following studies are indicated in gaucher disease:

  • Enzyme activity testing: Diagnosis can be confirmed through measurement of Glucocerebrosidase activity in peripheral blood leukocytes. A finding of less than 15% of mean normal activity is diagnostic.
  • Genotype testing: Molecular diagnosis can be helpful, especially in Ashkenazi patients, in whom 6 GBA mutations account for most disease alleles.
  • Radiography
    • Skeletal radiography can be used to detect and evaluate skeletal manifestations of gaucher disease.
  • Bone marrow examination for the presence of Gaucher’s cells is diagnostic.


 1) Enzyme replacement therapy (ERT) by recombinant β- Glucocerebrosidase is currently done. This preparation is highly effective in reversing the visceral and hematologic manifestations of gaucher disease.

2) Surgical Care

Partial and total Splenectomy was once advocated in the treatment of patients with gaucher disease. However, with the availability of ERT, this procedure is no longer necessary in most patients.

3)  Bone marrow transplant is also helpful.

4) Gene replacement is the permanent cure.


b) Prion disease              

The transmissible spongiform encephalopathies, or Prion diseases, are fatal neurodegenerative diseases characterized by spongiform changes, astrocytic gliomas, and neuronal loss resulting from the deposition of insoluble protein aggregates in neural cells.

They include Creutzfeldt-Jakob disease in humans, scrapie in sheep, and bovine spongiform encephalopathy (mad cow disease) in cattle.

Prion diseases may manifest themselves as infectious, genetic, or sporadic disorders. Because no viral or bacterial gene encoding the pathologic Prion protein could be identified, the source and mechanism of transmission of Prion disease long remained elusive.

It is recognized that Prion diseases are protein conformation diseases transmitted by altering the conformation, and hence the physical properties, of proteins endogenous to the host.

Human Prion-related protein, PrP, is monomeric and rich in α helix. Pathologic Prion proteins serve as the templates for the conformational transformation of normal PrP, known as PrPc, into PrPsc. PrPsc is rich in βsheet with many hydrophobic aminoacyl side chains exposed to solvent. PrPsc molecules therefore associate strongly with one other, forming insoluble protease-resistant aggregates. Since one pathologic Prion or Prion-related protein can serve as template for the conformational transformation of many times its number of PrPc molecules, Prion diseases can be transmitted by the protein alone without involvement of DNA or RNA.


c) Tay Sach disease

The GM2 Gangliosidosis include Tay-Sach’s disease that result from the deficiency of ß-Hexosaminidase activity and the lysosomal accumulation of GM2 gangliosides, particularly in the central nervous system, causing severe effects (neurodegeneration).

Inheritance– autosomal recessive trait having a predilection in the Ashkenazi Jewish population

Clinical symptoms and Classification

Tay-Sach’s disease is classified in variant forms, based on the time of onset of neurological symptoms. The variant forms reflect diversity in the mutation base.

  • Infantile TSD patients with this disease are born normal, but they develop loss of motor skills, increased startle reaction, macular pallor and retinal cherry red spot. Affected children develop normally till the age of 5-6 months, then decreased eye contact, hyperacusis (Exaggerated startle response) to noise are noted. Progressive development of idiocy and blindness are diagnostic of this disease and they are due to wide spread injury to ganglion cells, in brain and retina. The cherry red spot about the macula is due to destruction of retinal ganglion cells exposing the underlying vasculature.
  • Juvenile TSD. Extremely rare, Juvenile Tay-Sach’s disease usually presents itself in children between 2 and 10 years of age. They develop cognitive, motor, speech difficulties (dysarthria), swallowing difficulties (dysphagia), unsteadiness of gait (ataxia), and spasticity. Patients with Juvenile TSD usually die between 5–15 years.
  • Adult/Late Onset TSD. A rare form of the disorder, known as Adult Onset Tay-Sach’s disease or Late Onset Tay-Sachs disease (LOTS), occurs in patients in their 20s and early 30s. LOTS is frequently misdiagnosed, and is usually non-fatal. It is characterized by unsteadiness of gait and progressive neurological deterioration. Symptoms of LOTS, which present in adolescence or early adulthood, include speech and swallowing difficulties, unsteadiness of gait, spasticity, cognitive decline, and psychiatric illness, particularly schizophrenic-like psychosis


The diagnosis of infantile Tay-Sach’s disease is usually suspected in an infant with neurologic features and a cherry-red spot.

Enzymatic Assays-Definitive diagnosis is by determination of the level of ß-hexosaminidase in isolated blood leukocytes.


No cure for this disease. Symptomatic treatment is given. Enzyme replacement therapy and Gene therapy are under trial. Although experimental work is underway, no current medical treatment exists for infantile TSD. Patients receive palliative care to ease the symptoms. Infants are given feeding tubes when they can no longer swallow.


Prognosis is bad and death occurs in early years of life.

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