Disturbances of Aminoacid Metabolism


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      Dietary aminoacids (derived from the digestion of proteins) are deaminated and their amino group is disposed of in the urea cycle. Each aminoacid has its own catabolic pathway that allows the organism to derive energy from it. Aminoacids that can be converted to glycolytic intermediates and metabolyzed using the pathways of sugars are called glycogenic (in red in the figure below); aminoacids which can be converted to lipid-like products or acetil-CoA are called lipogenic (in blue in the figure below). Some aminoacids are split in two metabolytes, one lipogenic, the other glycogenic. The metabolism of Met, Ile and Val yields propionyl-CoA, which is common with the metabolism of uneven carbon fatty acids, but is a glycogenic metabolyte given that it is converted to succinyl-CoA and, via the Krebs cycle, to oxalacetic acid. When metabolytes of the Krebs cycle are produced, these are converted to oxalacetic acid, and this, in turn, is converted to piruvic acid for energy production. Several genetically determined defects of the metabolism of aminoacids are known some of them causing severe symptoms.

The fuel of the Krebs cycle is acetyl-CoA; thus reaching the intermediates of the Krebs cycle does not really ends the metabolic journey of aminoacids: it is necessary to convert oxaloacetate to acetil-CoA. This is accomplished by the enzyme P-enol piruvate carboxykinase which converts oxaloacetate to P-enolpiruvate, a glycolysis intermediate, from which piruvate can be obtained and, via piruvate dehydrogenase, acetyl-CoA.

      Some of the considerations given for the defects of the urea cycle also apply to defects of aminoacid metabolism. We add that: (i) in several cases, defects of aminoacid metabolism lead to overproduction of soluble metabolic intermediates that the kidney may eliminate; this reduces the severity of the disease. (ii) Careful dietary prescriptions may alleviate the symptoms by reducing the load of the aminoacid whose metabolism is impaired.
      The symptoms of hereditary defects of aminoacid metabolism are congenital or appear very early after birth, but contrary to the defects of the urea cycle, they are very variable; thus these are pediatric diagnoses. Mental retardation, possibly accompanied by seizures, is common to many of these defects; metabolic acidosis with increased anion gap (HAGMA), and/or Fanconi's syndrome (impaired reabsorption of solutes in the renal proximal tubuli, with glycosuria in the absence of hyperglycemia, phosphaturia, and loss of aminoacids and bicarbonate) occur in some of them. Some of these defects are mild or subclinical (e.g. alkaptonuria), others may cause extremely peculiar symptoms (e.g. albinism; urine smell like maple syrup in maple syrup urine disease). Diagnosis requires identification of the biochemical anomaly, and is confirmed by gene sequencing; aminoacid or their abnormal metabolytes are present in the blood and, at higher concentration, in the urine, which in many of these diseases is the biological sample of choice.
      9 aminoacids are essential (i.e. they need to be introduced with the diet, because humans do not have the enzymes required for their biosynthesis: phenylalanine (Phe), valine (Val), threonine (Thr), tryptophan (Trp), methionine (Met), leucine (Leu), isoleucine (Ile), lysine (Lys), and histidine (His). Genetic diseases in these cases are possible only for their catabolic pathways. Deficiency syndromes are described if their availability in the diet is insufficient.
      6 aminoacids are conditionally essential: arginine (Arg), cysteine (Cys), glycine (Gly), glutamine (Gln), proline (Pro), and tyrosine (Tyr). For these we have biosynthetic pathways but their biosynthesis may not be sufficient for our requirements or may depend on poorly available precursors (e.g. Tyr is produced from Phe; Cys from Met), or their decradation may be too effective (Arg). For these we have both biosynthetic and degradation pathways, and both can be affected by genetic diseases.
      5 aminoacids are non essential: alanine (Ala), aspartic acid (Asp), asparagine (Asn), glutamic acid (Glu), serine (Ser). These are produced from widely available intermediates of glycolysis or the Krebs cycle. Given their proximity to major metabolic pathways these are uusually not affected by genetic diseases.

Audio: Hereditary defects of aminoacid metabolism

Hereditary defects of aminoacid metabolism
Disease name Enzymatic defect Organs affected and symptoms Laboratory findings
Tyrosine and Phenylalanine
Phenylketonuria Hereditary deficiency of Phenylalanine hydroxylase (autosomal, recessive)Mental retardation (can be prevented by a diet poor in phenylalanine) Phenylpiruvic acid and other phenylketones in the urine; excess Phe in the blood
Tyrosinosis (Medes) Deficiency of Tyrosine alpha-ketoglutarate transaminase Mental retardation of variable severity excess Tyr in the blood
Tyrosinemia Deficiency of Fumarylacetoacetate hydrolase Fanconi's syndrome; liver failure excess Tyr in the blood
Alkaptonuria deficiency of Homogentisic oxidase benign (urine exposed to oxygen turns black) homogentisic acid in the urine
Albinism deficiency of Tyrosinase Absent melanine; hypopigmentation of the skin
Histidinemia (classic) Histidine ammonia lyase (liver and skin) Mental retardation of variable severity excess His in the blood
Histidinemia (variant) Histidine ammonia lyase (liver) Neurological damage; convulsive cryses; possible coma and death excess His in the blood
Branched chain aminoacidis (Valine, Leucine, and Isoleucine)
Maple syrup urine Disease Branched chain ketoacid decarboxylase Faticability; muscular cramps on exercise metabolic ketoacidosis: branched chain ketoacids in the serum
Valinemia deficiency of Valine aminotransferase Mental retardation of variable severity excess Val in the blood
Methylmalonic acidemia deficiency of Methylmalonyl-CoA mutase Mental retardation, acidosis and coma metabolic acidosis due to methylmalonic acid in the blood (and in the urine)
Sulfur containing aminoacids: Cysteine and Methionine
Homocystinemia deficiency of Cystathionine synthetase Skeletal abnormalities, mental retardation, thrombosis presence of homocysteine in the blood
Cystathioninemia deficiency of Cystathionase (Cystathionine Lyase) Skeletal abnormalities, mental retardation, thrombosis Cystathionine in the blood
Glycine and beta-alanine
Glycinemia deficiency of enzymes of the glycine degradation pathway Convulsive disease, mental retardation excess Gly in the blood
beta-alaninemia deficiency of beta-alanine aminotransferase Convulsive disease, possible coma and death presence of beta-Ala in the blood
Proline and hydroxyproline
Prolinemia, tipe I deficiency of Proline oxidase Nephritis (benign) excess Pro in the blood
Prolinemia, tipe II deficiency of Pyrroline 5-carboxylate dehydrogenase Mental retardation, convulsive disease excess Pro in the blood
Hydroxyprolinemia deficiency of Hydroxyproline oxidase Mental retardation presence of hydroxy-Pro in the blood
Lysinemia deficiency of Lysine ketoglutarate reductase Mental retardation; muscle weakness excess Lys in the blood
Saccharopinuria deficiency of saccharopine dehydrogenase (aminoadipic semialdheyde glutamate reductase; saccharopine pathway) Mental retardation Saccharopine in the blood and urine
Lysine intolerance deficiency of Lysine NAD oxidoreductase (pipecolate pathway) Vomiting, coma
Non proteic aminoacids
Ornithinemia Deficiency of Ornithine transaminase Athrophy of the choroid and retina Ornithine in the blood
Genetically determined disturbances of aminoacid transport
Hartnup diseasemembrane transporter for monoaminomonocarboxylic aminoacidsNeurological symptoms, vitamin B6 deficiencyTrp, Phe, Met, and other aminoacids in the urine
Cystinosis or cystinuria   Nephrolythiasis, renal insufficiency, Fanconi's syndrome presence of cystine (disulfide bond Cys dimer) in the urine

Audio: Metabolic acidosis and defects of aminoacid metabolism

      Laboratory diagnosis of hereditary defects of aminoacid metabolism is very important because dietary restrictions may mitigate or avoid mental retardation and other symptoms. As usual the main problem is suspecting the disease and asking the laboratory for the appropriate test. As a general rule, an aminoacid or its metabolyte will be present in the serum and urine; and since the differential diagnosis cannot be effected on clinical grounds, the different diseases often resembling each other, it is reasonable to measure all the possible metabolytes listed in the table. A possible association that may call your attention is mental retardation and metabolic acidosis. Once the aminoacid or metabolyte has been identified, and a diagnosis is suspected, confirmation may be obtained by means of gene sequencing.


      Some aminoacids have so simple metabolic pathways that genetic diseases are virtually unknown; e.g. alanine is transaminated to piruvate in a single reaction step. Other aminoacids may have complex metabolic pathways requiring several enzymes, and prone to disease.

GLYCINE. In spite of its simple formula and its different possible catabolic pathways, diseases due to genetic defects of Gly catabolism are described. It is interesting to point out that one of Gly catabolic pathways produces oxalic acid, which is excreted by the kidney and may precipitate in the form of calcium oxalate forming urinary stones. Thus imbalances in Gly metabolism (inherited or acquired), may be associated to urolythiasis.

Ala, Ser, Glu, Gln, Asp, Asn are deaminated by single enzymes to produce piruvate, β-ketoglutarate or oxaloacetate. These are metabolytes of the Krebs cycle or glycolysis. Since these pathways are always or almost always functional (and these aminoacids are water-soluble), confirmed diseases of the metabolism of the above aminoacids essentially do not occur.

CYSTEINE. Cysteine is metabolized via several pathways, the principal of which are depicted below. Sulfur is eliminated as sulfite or sulfate. Taurine plays several physiological roles (e.g. in the stabilization of the lipid bilayers) and can be eliminated in the bile and in the urine.

PHENYLALANINE AND TYROSINE. Phenylalanine(Phe) and tyrosine (Tyr) can be interconverted in a reaction catalyzed by phenylalanine hydroxylase, thus they share common metabolic pathways, and the relative disturbances. Phe and Tyr are essential aminoacids, for which humans do not possess biosynthetic pathways. Their catabolism is specially complex because besides their degradation pathway, leading to fumarate and acetoacetate, they are the precursors of catecholamine hormones (dopamine, adrenaline and noradrenaline), and of melanins the skin pigments responsible for the color of our skin and hair.
      The most frequent defect of aminoacid catabolism is phenylketonuria that is due to an autosomal recessive deficiency of Phe-hydroxylase. Lack of this enzyme impairs the conversion of Phe to Tyr, which is the first step of Phe catabolism. Phe accumulates in the blood and the urine and is converted to phenylpiruvic acid and its metabolytes phenyllactate and phenylacetate, which also appear in the blood and urine. Early diagnosis is essential because the baby is normal at birth (the mother provides to metabolyze Phe in excess) and a diet artifically deprived of Phe may prevent the severe mental retardation associated to accumulation of phenylketones in the brain. An important observation is the following: Phe-hydroxylase requires biopterin as a redox cofactor. . Biopterin has a structure similar to folate but is not a vitamin, and can be synthesized starting from GTP and the enzymes required for its biosynthesis may also be affected by genetic defects. Genetic defects of the biosynthesis of biopterin may mimic phenylketonuria, but the cofactor is also involved in other metabolic pathways, thus accumulation of Phe is not the only symptom in these cases, and the treatments is better and more simply carried out by administration of dietary biopterin supplements.

      The catabolism of Phe and Tyr leads to fumarate and acetoacetate; important intermediates are p-hydroxyphenyl piruvate and homogentisate. The metabolic pathway is depicted below (in black the enzymes catalyzing each reaction step; in red the genetic disease due to their deficiency):

      The biosynthesis of melanins, the pigments responsible fo the color of our skin, iris and hair starts from the oxidation of Tyr by the enzyme tyrosinase. A genetic defect of tyrosinase causes albinism, the lack of skin pigments.

HISTIDINE. His is converted to glutamic acid in a short metabolic pathway whose first step of this pathway is catalyzed by the enzyme His ammonia lyase and produces one molecule of ammonia. At keast two different genetic defects of this enzyme are described and cause variants of the severe inherited disease histidinemia.

PROLINE and ARGININE are converted to glutamic acid. One genetic disease is described for the catabolism of Pro, prolinemia, due to the defect of proline oxidase, and two diseases are described for the catabolism of Arg, argininemia and ornithinemia. Arginase, the enzyme whose defect causes argininemia belongs to the urea cycle and was described together with other genetic defects of that cycle.

BRANCHED CHAIN AMINOACIDS and PROPIONYL-CoA. Branched chain aminoacids (Val, Leu, Ile) have a partly common pathway whose most important inherited defect are valinemia, due to the absence of the relative transaminase, and maple syrup urine disease, due to the absence of branched chain ketoacid decarboxylase:

Moreover, the catabolism of branched chain amnoacids is among the main producers of propionyl-CoA, a metabolyte common also to the catabolism of uneven C fatty acids and other pathways. A defect of the enxyme methylmalonyl-CoA mutase or of the metabolism of vitamin B12 (required as a cofactor of methylmalonyl-CoA mutase), cause accumulation of methylmalonyl-CoA which spontaneously looses CoA-SH and is converted to methylmalonic acid. Methylmalonic acid is excreted in the urine, but its accumulation in the blood causes a metabolic acidosis (methylmalonic acidemia or aciduria). This metabolyte is easily identified in the serum or in the urine by the clinical laboratory.

METHIONINE. Met is an essential aminoacid which plays two major functions: is a constituent of proteins, and a constituent of the coenzyme S-adenosyl-methionine (SAM), a methyl group donor. The catabolism of Metis complex because: (i) it requires conersion to SAM; (ii) it is partially reversible (i.e. one of its products homocysteine can be converted back to Met); (iii) it is also the biosynthetic pathway of Cys.
      There are two important inherited disease of Met catabolism, which cause an increase in the plasma concentration of homocysteine and cystathionine. Homocysteinemia is due to a defect of the enzyme cystathionine synthase. Two variants of the defect are known, one of which can be treated with administration of high doses of vitamin B6 (pyridoxine), the cofactor of cystathionine synthase. Homocysteinemia may cause mental retardation and very early atherosclerosi. Moreover, sulfur containing aminoacids present in excess in the serum favor platelet aggregation and thrombosis. The laboratory diagnosis relies upon the finding of high concentrations of homocysteine in the blood and in the urine. Cystathioninemia is due to the defect of cystathionine lyase and causes an increase of cystathionine in the plasma and in the urine.

LYSINE. The catabolism of the essential aminoacid lysine is extremely complex. Two main pathways are described, both of which can be affected by inheritable diseases: the saccharopine pathway, occurring partly in the cytoplasm and partly in the mitochondria, and the pipecolate pathway, occurring partly in the cytoplasm and partly in the peroxisomes. The two pathways converge at the common intermediate α-aminoadipic semialdehyde. Conversion of Lys to α-aminoadipic semialdehyde replaces an amino group with a carbonyl and is formally equivalent to an oxidative transamination or the oxidation, but requires a more complex pathway because the amino group in ε is less reactive than in α. Many organs carry out Lys catabolism, but the principal one is the liver, which mainly uses the saccharopine pathway. The brain is atypical in that it mainly uses the pipecolate pathway.

TRYPTOPHAN. The main catabolic pathway of the essential aminoacid Trp is depicted below:

      A very important feature of Trp catabolism is that it provides part of our daily requirement of vitamin B3 (nicotinamide). The vitamin thus produced does not suffice to make us independent of its external supply, but in the presence of: (i) a defect of Trp catabolism; or (ii) an insufficient dietary apport of this aminoacid; or (iii) a loss of it (as it occurs in Hartnup disease) the daily requirement of nicotinamide is significantly increased. A sepcific avitaminosis (pellagra) may result.
      The most important genetic defect of Trp catabolism is xanthurenic aciduria due to a deficiency of the enzyme kynureninase. This disease is rare and may cause mental retardation if untreated. Kinureninase uses PLP as a cofactor and two variants of xanthurenic aciduria are known: a pyridoxine sensitive and a pyridoxine insensitive one. The most likely explanation is as follows: mutations causing reduced affinity of the enzyme for PLP may respond to high doses of the cofactor; mutations that inactivate the enzyme or prevent its biosynthesis are unresponsive.

α-KETOADIPIC ACID is a metabolyte encountered in the catabolic pathways of Lys and Trp. The conversion of α-ketoadipic acid to acetyl-Coa requires two specialized enzymes, α-ketoadipic acid dehydrogenase, which decarboxylates the substrate and combines it with CoA-SH, in a reaction which is reminiscent of those catalyzed by piruvate dehydrogenase or α-ketoglutarate dehydrogenase, and glutaryl-CoA dehydrogenase. The product of the two sequential reactions catalyzed by these enzymes is crotonyl-CoA, which can enter the last steps of the β-oxidation pathway.

Two clinically similar genetic defects may affect the metabolism of α-ketoadipic acid: ketoadipic aciduria, and glutaric aciduria. The main symptom of both is mental retardation. The peculiarity of these diseases is that two aminoacids (Trp and Lys) are involved in their determinism, and dietary restrictions apply to both of them.


      Hartnup disease strictly speaking is not a defect of aminoacid metabolism, but a defect of their absorption in the gut and in the renal tubuli. It is inherited as an autosomal recessive trait affecting a membrane transporter for monoaminomonocarboxylic aminoacids. Tryptophan, phenilalanine, methionine and other aminoacids appear in the urine. The defect of tryptophan absorption in the gut is specially relevant, because this aminoacid, besides being essential, is the precursor of vitamin B3 (niacine, the precursor of NADH and NADPH). As a consequence, in Hartnup disease one may observe the cutaneous and neurological symptoms of vitamin B3 deficiency (pellagra). Diagnosis is based on the identification of multiple aminoacids in the urine, together with triptophan metabolytes. The prognosis is good provided that an adequate dietary regimen is instituted.

      Cystinuria is a defect of aminoacid transport, conceptually similar to Hartnup disease. The aminoacids that appear in the urine are lysine, arginine, ornithine and cystine (the dimer of cysteine). Cysteine, being poorly soluble precipitates yielding yellow calculi, that strongly suggest the diagnosis; confirmation comes from the demonstration of above-mentioned aminoacids in the urine. Cystinuria is inherited as an autosomal recessive trait and is relatively benign, nephrolythiasis being the most important symptom; however on the long run renal insufficiency may develop.

      Fanconi's syndrome and cystinosis. These two conditions are often associated. Cystinosis causes an accumulation of Cys in several tissues. Fanconi's syndrome, which may be inherited or acquired is a defective renal reabsorption of several aminoacids, glucose, phosphate and bicarbonate. Loss of bicarbonate causes a metabolic acidosis. Fanconi's syndrome may be inherited or acquired. The inherited form is an autosomal recessive traits and is associated to cystinosis (presumably because of a defective transport of Cys in other tissues besides the proximal tubuli). The acquired form is usually secondary to toxic or neoplastic damage of the kidney.

Audio: Hereditary defects of aminoacid transport


      The diagnosis of inherited defects of the metabolism of aminoacids is difficult. Each singe disease is rare, but since these diseases are many, their cumulative incidence is significant, probably in the order of 2-4 per one thousand births. All these diseases manifest themselves early in extrauterine life, thus diagnosis is usually called for in the first months of life. The correct diagnosis is important because the baby is usually normal at birth, and specific dietary restriction may in some cases prevent brain damage.
      The urine is an ideal sample for broad scope screening of this type of metabolic defects because it concentrates the aminoacid or metabolyte, and because the normal urine contains very few organic compounds: urea, creatinine, bicarbonate, some bilirubin and little else. A good starting point is the analysis of organic compounds in the baby's urine by thin layer chromatography. To carry out this analysis a drop of the patient's urine is adsorbed on chromatographic paper or alluminium oxide plate, and the medium is inserted in the chosen solvent (water or water/ethanol for paper, organic solvents for alumina). The solvent is allowed to flow over the medium by capillary absorption, then the medium is stained for organic compounds. The different organic compounds will move from the initial position, more or less according to their solubility in the chosen solvent. A standard sample containing the most common aminoacids and metabolytes may be run in parallel to help the identification of the analytes.

      Babies presenting somnolescence, stunted growth, seizures, etc. should be investigated for inherited metabolic defects. Familial anamnesis may offer some clue, but is often negative, these diseases being inherited as recessive traits.

      The determination of the concentration of the 20 aminoacids in the serum and the urine by means of high performance liquid chromatography and mass spectrometry may be the next step. This exam may reveal the abnormal aminoacid, or may reveal the presence of an abnormal compound, not corresponding to any aminoacid, but possibly indicative of one of their metabolytes.

HPL chromatography of urine aminoacids, detected by conductance. Marked increases of Trp is evident.

      The screening of the most common of these diseases, phenylketonuria is routinely carried out at birth by measuring the concentration of phenylpiruvic acid in the urine. A hemogas analysis may reveal a metabolic acidosis with increased anion gap, indicative of the presence of an acidic metabolyte in the serum.
      Once the presence of an abnormal metabolyte in the urine or in the serum has been detected, it can usually be identified by chemical or enzymatic methods. The diagnostic suspicion should be confirmed by gene sequencing. It is advisable to submit to genetic sequencing also the baby's parents, for genetic counseling.

Further readings
BK Burton Inborn Errors of Metabolism in Infancy: A Guide to Diagnosis. Pediatrics 1998; 102: (6) e69.
J Häberle, A Chakrapani, N Ah Mew, and N Longo Hyperammonaemia in classic organic acidaemias: a review of the literature and two case histories
SC Sreenath Nagamani, A Erez, and B Lee Argininosuccinate Lyase Deficiency.
Andre' A. and Jamie J.F. Lysine metabolism in mammalian brain: an update on the importance of recent discoveries . Amino Acids 2013, 45: 1249-72.
Patil V.S. et al. Screening for aminoacidurias and organic acidurias in patients with metabolic or neurological manifestations Biomedical Research 2012; 2: 253-258,

Questions and exercises:
1) Hyperammoniemia suggests:
An hereditary defect in aminoacid metabolism
An hereditary defect of the urea cycle
An hereditary defect of aminoacid transport

2) Phenylketonuria is among the most common hereditary defects of metabolism; the aminoacid whose metabolism is impaired is/are:

3) Inherited defects of aminoacid metabolism may cause metabolic acidosis. In these cases the anion gap is

4) Albinism causes a severe photosensitivity of the skin because of the lack of the protection offered by melanins. An important differential diagnosis is:
Porphyria erytropoietica congenita
Crigler Najjar syndrome
Lesch-Nyhan syndrome

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Thank you Professor (lecture on bilirubin and jaundice).

The fourth recorded part, the one on hyper and hypoglycemias is not working.
Bellelli: I checked and in my computer it seems to work. Can you better specify
the problem you observe?

This Presentation (electrolytes and blood pH) feels longer than previous lectures
Bellelli: it is indeed. Some subjects require more information than others. I was
thinking of splitting it in two nest year.

Bellelli in response to a question raised by email: when we compare the blood pH
with the standard pH we do not mean to compare the "normal" blood pH (7.4)
with the standard pH. Rather we compare the actual blood pH of the patient, with
the pH of the same blood sample equilibrated under standard conditions.
Thus, if we say that standard pH is lower than pH we mean that equilibriation with
40 mmHg CO2 has caused absorption of CO2 and has lowered the pH with respect
to its value before equilibration.

(Lipoproteins) Is the production of leptin an indirect cause of type 2 diabetes since
it works as a stimulus to have more adipose tissue that produces hormones?
Bellelli: in a sense yes, sustained increase of leptin causes the hypothalamus to adapt
and to stop responding. Obesity ensues and this in turn may cause an increase in the
production of resistin and other insulin-suppressing protein hormones produced by the
adipose tissue. However, this is quite an indirect link, and most probably other factors
contribute as well.

(Urea cycle) what is the meaning of "dissimilatory pathway"?
Bellelli: a dissimilatory pathway is a catabolic pathway whose function is not to produce
energy, but to produce some terminal metabolyte that must be excreted. Dissimilatory
pathways are necessary for those metabolytes that cannot be excreted as such by the
kidney or the liver because they are toxic or poorly soluble. Examples of metabolytes
that require transformation before being eliminated are heme-bilirubin, ammonia,
sulfur and nitrogen oxides, etc.

Talking about IDDM linked neuropathy can be the C peptide absence considered a cause of it??
Bellelli: The C peptide released during the maturation of insulin, besides being an indicator
of the severity of diabetes, plays some incompletely understood physiological roles. For
example it has been hypothesized that it may play a role in the reparation of the
atherosclerotic damage of the small arteries. Thus said, I am not aware that it plays a direct
role in preventing diabetic polyneuropathy. Diabetic neuropathy has at least two causes: the
microvascular damage of the arteries of the nerve (the vasa nervorum), and a direct
effect of hyperglycemia and decreased and irregular insulin supply on the nerve metabolism.
Diabetic neuropathy is observed in both IDDM and NIDDM, and requires several years to
develop. Since the levels of the C peptide differ in IDDM and NIDDM, this would suggest
that the role of the C peptide in diabetic neuropathy is not a major one. If you do have
better information please share it on this site!

In acute intermitted porphyria and congenital erythropoietic porphyria why do the end product
of the affected enzymes accumulate instead of their substrate??
Bellelli: First of all, congratulations! This is an excellent question.
Remember that a condition is which the heme is not produced is lethal in the foetus; thus
the affected enzyme(s) must maintain some functionality for the patient
to be born and to come to medical attention. All known genetic defects of heme
biosynthesis derange but do not block this metabolic pathway.
Congenital Erythropoietc Porphyria (CEP) is a genetic defect of uroporphyrinogen
III cosynthase. This protein associates to uroporphyrinogen synthase (which is present
and functional in CEP) and guarantees that the appropriate uroporphyrinogen isomer is produced
(i.e. uroporphyrinogen III). In the absence of a functional uroporphyrinogen III
cosynthase other possible isomers of uroporphyrinogen are produced together with
uroporpyrinogen III, mostly uroporphyrinogen I. The isomers of uroporphyrinogen
that are produced differ because of the positions of propionate and acetate side chains,
and this in turn is due to the pseudo symmetric structure of porphobilinogen. Only
isomer III can be further used to produce protoporphyrin IX. Thus in the
case of CEP we observe accumulation of abnormal uroporphyrinogen derivatives, which, as
you correctly observed are the products of the enzymatic synthesis operated by
uroporphyrinogen synthase.
The case of Acute Intermittent Porphyria (AIP) is similar, although there may be variants
of this disease. What happens is that either the affected enzyme is a variant that does not
properly associate with uroporphyrinogen III cosynthase or presents active site mutations
that impair the proper alignement of the phoprphobilinogen substrates. In either case
abnormal isomers of uroporphyrinogen are produced, as in CEP.
Also remark that in both AIP and CEP we observe accumulation of the porphobilinogen
precursor: this is because the overall efficiency of the biosynthesis of uroporphyrinogens is
reduced. Thus: (i) less uroporphyrinogen is produced, and (ii) only a fraction of the
uroporphyrinogen that is produced is the correct isomer (uroporphyrinogen III).

is it possible to take gulonolactone oxidase to synthesize vitamin C
instead of vitamin C supplement?
Bellelli: no, this approach does not work. The main reason is that
the biosynthesis of vitamin C, as almost all other metabolic processes, occurs intracellularly.
If you administer the enzyme it will at most reach the extracellular fluid but will not be
transported inside the cells to any significant extent. Besides, there are other problems
in this type of therapy (e.g. the enzyme if administered orally, may be degraded by digestive
proteases; if administered parenterally, may cause the immune system to react against a
non-self protein). In theory one could think of a genetic modification of the inactive human
gene of gulonolactone oxidase, but the risk and cost of this intervention would not be
justified. In addition to these considerations, except for cases of shipwreckage or
other catastrophes, a proper diet or administration of tablets of vitamin C is effective,
risk-free and unexpensive, thus no alternative therapy is reasonable. However, I express my
congratulations for your search on the biosynthesis pathway of ascorbic acid.

Resorption and not reabsorption would lead to hypercalcemia ie bone matrix being broken down.
Bellelli: I am not sure to interpret your question correctly. Resorption indicates destruction of the bone matrix and release of calcium and
phosphate in the blood, thus it causes an increase of calcemia. Reabsorption usually means active transport of calcium from the renal tubuli to the blood, thus
it prevents calcium loss. It prevents hypocalcemia, and thus complement bone resorption. To avoid confusion it is better use the terms "bone resorption" and "
renal reabsorption of calcium". If you have a defect in renal reabsorption, parthyroid hormone will be released to maintain a normal calcium level by means of
bone resorption; the drawback is osteoporosis.

In Reed and Frost model: I haven't understood what is the relationship
between K and R reproductive index. Thank you Professor!
Bellelli: in the Reed and Frost model K is the theoretical upper limit of
R0. R the reproductive index is the ratio (new cases)/(old cases) measured after
one serial generation time. R0 is the value of R one measures at the beginning
of the epidemics, when in principle all the population is susceptible.

What is the link between nucleotide metabolism and immunodeficiencies and mental retardation?
Bellelli: the links may be quite complex, but the principal ones are as follows:
1) the immune response requires a replication burst of granulocytes and lymphocytes, which in turn requires
a sudden increase of nucleotide production, necessary for DNA replication. Defects of nucleotide metabolism
impair this phase of the immune defense. Notice that the mechanism is similar to the one responsible of
anemia which requires a sustained biosynthesis of nucleotides at a constant rate, rather than in a burst.
2) Mental retardation is mainly due to the accumulation of nulceotide precursors in the brain of the
newborn, due to the incompletely competent blood-brain barrier.

How can ornithine transaminase defects cause hyperammonemia? Is it due to the accumulation
of ornithine that blocks the urea cycle or for other reasons?
Bellelli: ornithine transaminase is required for the reversible interconversion of ornithine
and proline, and thus participates to both the biosynthesis and degradation of ornithine. The enzyme is
synthesized in the cytoplasm and imported in the mitochondrion. Depending on the metabolic conditions
the deficiency of this enzyme may cause both excess (when degradation would be necessary) or defect
(when biosynthesis would be necessary) of ornithine; in the latter case, the urea cycle slows down. Thus
there is the paradoxical condition in which alternation may occur between episodes of hyperammonemia
and of hyperornithinemia.

When we use the Berthelot's reaction to measure BUN do we also have to
measure the concentration of free ammonia before adding urease?
Bellelli: yes, in principle you should. Berthelot's reaction detects ammonia,
thus one should take two identical volumes of serum, use one to measure free ammonia,
the other to add urease and measure free ammonia plus ammonia released by urea. BUN is
obtained by difference. However, free ammonia in our blood is so much lower than urea that
you may omit the first sample, if you only want to measure BUN.

Why do we have abnormal electrolytes in hematological neoplasia e.g.
Bellelli: I do not have a good explanation for this effect, which may have
multiple causes. However, you should consider two factors: (i) acute leukemias cause a massive
proliferation of leukocytes (or lymphocytes depending on the cell type affected) with a very
shortened lifetime; thus you observe an excess death rate of the neoplastic cells. The dying
cells release in the bloodstream their content, which has an electrolyte composition different
from that of plasma: the cell cytoplasm is rich in K and poor in Na, thus causing hyperkalemia.
(ii) the kidney may be affected by the accumulation of neoplastic white cells or their lytic products.

Gaussian curve: If it is bimodal is it more likely to be a "certain diagnosis" than if it is
unimodal or does it only show the distinguishment from health?
Bellelli an obviously bimodal Gaussian curve indicates that the disease is clearly
separated from health: usually it is a matter of how precise and clear-cut is the definition of the disease.
For example tuberculosis is the disease caused by M. tuberculosis, thus if the culture of the sputum is
positive for this bacterium you have a "certain" diagnosis (caution: the patient may suffer of two diseases,
e.g. tuberculosis and COPD diagnosis of the first does not exclude the second). However, in order to have
a "certain" diagnosis it is not enough that distribution of the parameter is bimodal, it is also required that the
patient's parameter is out of the range of the healthy condition: this is because a distribution can be
bimodal even though it is composed by two Gaussians that present a large overlap, and the patient's
parameter may fall in the overlapping region. Thus, in order to obtain a "certain" diagnosis you need to
consider not only the distribution of the parameter(s) but also the patient's values and the extent of the
overlapping region.

Prof can you please elaborate a bit more on the interhuman variability and its difference
with the interpopulation variability please?
Bellelli: every individual is a unique combination of different alleles of the same genes;
this is the source of interindividual variability. Every population is a group of individuals who intermarry and
share the same gene pool (better: allele pool). Every allele in a population has its own frequency. Two
population may differ because of the diffferent frequencies of the same alleles; in some cases one
population may completely lack some alleles. The number and frequencies of alleles of each gene
determine the variance. If you take two populations and calculate the cumulative interindividual variance
of the population the number you obtain is the sum of two contributions: the interindividual variance within each population, plus the interpopulation variance
between the means of the allele frequencies. For example, there are human population in which the frequency of blood group B is close to 0% and other populati
ons in which it is 30% or more.

Prof can you please explain again the graph you have showed us in class about thromboplastin?
(Y axis=abs X axis= time)
Bellelli: the graph that I crudely sketched in class represented the signal
of the instrument (an absorbance spectrophotometer) used to record the turbidity of the
sample (turbidimetry). The plasma is more or less transparent, before coagulation starts.
When calcium and the tissue factor (or collagen) are added. thrombin is activated and begins
digesting fibrinogen to fibrin; then fibrin aggregates. The macroscopic fibrin aggregates cause
the sample to become turbid, which means it scatters the incident light. The instrument reads
this as a decrease of transmitted light ( an increase of the apparent absorbance) and the
time profile of the signal presents an initial lag phase, which is called the protrombin or
thromboplastin time depending on the component which was added to start coagulation
(tissue factor or collagen).

Prof can you please explain the concept you have described in class about
the simultaneous hypercoagulation and hemorrhagic syndrome? How can this occur?
Bellelli: The condition you describe is observed only in the Disseminated
Intravascular Coagulation syndrome. Suppose that the patient experiences an episode of
acute pancreatitis: tripsin and chymotripsin are reabsorbed in the blood and proteolytically
activate coagulation causing an extensive consumption of fibrinogen and other coagulation
factors. Tripsin and chymotripsin also damage the vessel walls and may cause internal
hemorrages, but at that point the consumption of fibrinogen may have been so massive that
not enough is left to form the clot where the vessel has been damaged, causing an internal
hemorrage. Pancreatitis is a very severe, potentially lethal condition, and DIC is only one of
the reasons of its severity.

You said that certain drugs (ethanol, cocaine, cannabis, opiates...) cause a
necessity of higher and higher dosage, for two reasons: the enzyme in the liver is inducible and
the receptors in the brain are expressed less and less. So, first, I am not sure I got it right, and
second I did not understand how expressing less receptors leads to a necessity of higher
Bellelli: You got it correctly, but the detailed mechanism of resistance may
vary among different substances, and not all drugs cause adaptation.
The reason why reducing the number of receptors may require an increased dosage of the drug
is as follows: suppose that a certain cell has 10,000 receptors for a drug. When bound to its
agonist/effector, each receptor produces an intracellular second messenger. Suppose that in
order for the cell to respond 1,000 receptors must be activated. The concentration of the
effector required is thus the concentration that produces 10% saturation. You can easily
calculate that this concentration is approximately 1/10 of the equilibrium dissociation constant
of the receptor-effector complex (its Kd), the law being
Fraction bound = [X] / ([X]+Kd)
where [X] is the concentration of the free drug.
After repeated administration, the subject becomes adapted to the drug, and his/her cells
express less receptors, say 5,000. The cell response will in any case require that 1,000
receptors are bound to the effector and activated, but this now represents 20% of the total
receptors, instead of 10%. The drug concentration required is now 1/4 of the Kd.
Continuing administration of the drug further reduces the cell receptors, but the absolute
number of activated receptors required to start the response is constant; thus the fewer
receptors on the cell membrane, the higher the fraction of activated receptors required.

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