Holocarboxylase Synthetase Deficiency

Updated: Apr 17, 2018
Author: Karl S Roth, MD; Chief Editor: Luis O Rohena, MD, PhD, FAAP, FACMG 



Holocarboxylase synthetase (HCS) deficiency was defined as a distinct genetic disorder several years after its initial clinical description, similar to the discovery of propionic acidemia. From 1970-1973, only 3 clinically varied cases of children who excreted beta-methylcrotonic acid in their urine were reported. Notable differences included the age at presentation (neonatal to 9 mo), presence of acidosis, and response to biotin administration.

In 1976, Roth et al reported an infant who was the second affected sibling born to an otherwise healthy mother. The first child, a male, died from aspiration and severe acidosis 36 hours after birth; no autopsy or other information was available. From birth, the second child was clinically affected by an unrelenting metabolic acidosis and a massive excretion of beta-methylcrotonic acid, lactic acid, beta-hydroxybutyric acid, and beta-hydroxypropionic acid. This child died 105 hours postdelivery after little improvement from small intravenous doses of megavitamins, including biotin.[1] Subsequent studies using fibroblasts recovered from the infant demonstrated a defect of carbon dioxide fixation into pyruvate, propionate, and beta-methylcrotonate; thus, the term multiple carboxylase deficiency was applied to the disorder.

Recognizing the unlikely coexistence of 3 gene mutations in a single individual, research continued in an attempt to find a unifying factor to explain the involvement of 3 distinct enzymes. Patients seemed to respond to biotin administration, and each affected enzyme required biotin as a cofactor. This directed attention toward the work of Lynen, who had previously demonstrated that carbon dioxide fixation in microbes required covalent attachment of the biotin to an apoprotein. Since covalent bond formation requires the mediation of an enzyme, a search began for an enzyme deficiency that was common to all 3 carboxylases and that might explain the defective function of each.

Finally, in 1980, based on the work of Saunders and coworkers, Roth et al reported holocarboxylase synthetase deficiency in a subsequent sibling of their original case; the sibling had an excellent clinical response to biotin administration in pharmacologic doses.[2, 3] In 1981, Burri et al reported evidence of holocarboxylase synthetase deficiency in this patient and in others with a similar neonatal presentation; thus, the nature of the defect was redefined as a single enzyme defect.[4] The late or juvenile-onset type of presentation, which resembled the newly defined neonatal holocarboxylase synthetase deficiency in virtually all respects other than age at onset, still required an explanation.

In 1983, Wolf et al suggested that the late-onset type may be due to a defect in biotin recycling rather than to defective absorption; others had proposed the same finding.[5] Recycling is well described for other vitamins, helping explain the normally miniscule daily requirements. Logically, the same should hold true for biotin.

The major contribution of Wolf et al was in demonstrating the presence of the enzyme human biotinidase and its role in biotin recycling. Late-onset multiple carboxylase deficiency is now known to be due to a biotinidase deficiency and subsequent impairment of biotin recycling. Thus, what was originally reported as the disease beta-methylcrotonic aciduria has been separated into 3 distinct genetic disorders: beta-methylcrotonyl-coenzyme A (CoA) carboxylase deficiency, holocarboxylase synthetase deficiency, and biotinidase deficiency. This article focuses on the specific defect recognized as holocarboxylase synthetase deficiency.


Carbon dioxide fixation, a process typically associated with plant metabolism, is a vital reaction in humans. In individuals with multiple carboxylase deficiency, more than 3 metabolic pathways are impaired. Carbon dioxide fixation occurs in the metabolism of several different substrates; thus, several pathways are involved in the defect. Acetyl-CoA carboxylase (ACC) is key to fatty acid synthesis, pyruvate carboxylase is a critical step in gluconeogenesis, propionyl-CoA carboxylase produces methylmalonyl-CoA prior to the ultimate formation of succinyl-CoA, and beta-methylcrotonyl-CoA carboxylase is critical in the degradation of leucine for energy. Thus, affected infants have increased levels of circulating odd-chain fatty acids (due to deficient propionyl-CoA carboxylase), hypoglycemia (deficient pyruvate carboxylase), and ketoacidosis (deficient propionyl-CoA carboxylase and beta-methylcrotonyl-CoA carboxylase).[6]

As in all other enzyme reactions that require a cofactor, normal production of each of the carboxylase apoproteins is insufficient to carry out their individual carbon dioxide–fixation reactions. Since all enzyme function requires binding between the substrate and enzyme, impairment of such binding prevents enzyme function. In apocarboxylases, the binding of each to carbon dioxide requires the presence of biotin, to which the carbon dioxide physically attaches.

For a carboxylase protein to function normally, a covalent bond with the cofactor biotin must be established. Because covalency involves a great deal of bond energy, such bonds require enzyme-mediated reactions before they can occur efficiently. The specific function of holocarboxylase synthetase is to establish a covalent bond between a lysine residue in the apocarboxylase molecule and a biotin molecule.[7, 8]

Therefore, holocarboxylase synthetase deficiency impairs all carbon dioxide–fixation reactions within the cell. These include reactions mediated by acetyl-CoA carboxylase, pyruvate carboxylase, propionyl-CoA carboxylase, and beta-methylcrotonyl-CoA carboxylase. Each of these serves an extremely important cellular function, and impairment has significant adverse consequences that manifest as clinical disease. A recent study reported that the liver and brain have different mechanisms of holocarboxylase synthetase gene regulation, suggesting that brain function is conserved at the expense of somatic cell function in biotin deficiency.

A study of holocarboxylase synthetase messenger RNA (mRNA) from 3 different human cell lines suggests the existence of 3 distinct types of mRNA originating from separate exons. Thus, many holocarboxylase synthetase mutations may remain undetected. A separate report of a partially responsive patient given extraordinarily high doses of biotin may represent one such example.

Acetyl-CoA carboxylase has an important cellular function as the first step in fatty acid synthesis. The 3-carbon fatty acyl compound malonyl-CoA is formed through carboxylation of acetyl-CoA. Regulation of acetyl-CoA carboxylase activity is important to the energy economy of the cell, since excess glucose is normally converted to lipid through this reaction. Although no individual symptomatology may be directly attributed to functional impairment, lactate accumulation in infants with holocarboxylase synthetase deficiency may be partially attributable to the inability to channel acetyl-CoA (produced from glucose oxidation) into fatty acid synthesis.

Pyruvate carboxylase is a major step in the process of gluconeogenesis, a sequence of reactions that results in resynthesis of a 6-carbon glucose molecule from two 3-carbon fragments produced chiefly from glycolysis. Interruption of gluconeogenesis is likely to result in clinical hypoglycemia; this is a well-known finding in both isolated pyruvate carboxylase deficiency and holocarboxylase synthetase deficiency. In addition, hypoglycemia encourages oxidation of fatty acids, including essential fatty acids, resulting in the development of a typical exfoliative dermatitis.

The substrates for both propionyl-CoA and beta-methylcrotonyl-CoA carboxylases are organic acids that contain an alpha-keto group; failure to fix carbon dioxide through these 2 enzyme reactions results in accumulation of keto-acids and clinical ketoacidosis, accompanied by hyperglycinemia and hyperammonemia.

In general, the organic acids inhibit the urea cycle, which diminishes incorporation of free ammonia. This effect is exerted at the site of N -acetylglutamate synthetase (NAGS), which mediates production of the activating substance (N -acetylglutamate [NAG]) for carbamyl phosphate synthetase, the first step in the urea cycle. Diminished activation slows ammonia incorporation, with accumulation in blood and other tissues. See the image below.

Urea cycle. Compounds that comprise the urea cycle Urea cycle. Compounds that comprise the urea cycle are numbered sequentially, beginning with carbamyl phosphate. At the first step (1), the first waste nitrogen is incorporated into the cycle; also at this step, N-acetylglutamate exerts its regulatory control on the mediating enzyme, carbamyl phosphate synthetase (CPS). Compound 2 is citrulline, the product of condensation between carbamyl phosphate (1) and ornithine (8); the mediating enzyme is ornithine transcarbamylase. Compound 3 is aspartic acid, which is combined with citrulline to form argininosuccinic acid (4); the reaction is mediated by argininosuccinate (ASA) synthetase. Compound 5 is fumaric acid generated in the reaction that converts ASA to arginine (6), which is mediated by ASA lyase.

Thus, the major clinical findings in individuals with holocarboxylase synthetase deficiency, described in detail by Roth and coworkers, include severe ketoacidosis, exfoliative dermatitis, and hypoglycemia.[3] The urine may have a distinctive tomcatlike odor; however, the odiferous constituent has not been characterized.



United States

The true incidence in the newborn population cannot be cited for lack of data based on population screening. Holocarboxylase synthetase deficiency is among the rarest of inborn errors, with an estimated incidence of less than 1 per 200,000 live births.


Without early diagnosis and treatment, the mortality rate is close to 100%.[9] With treatment, morbidity depends on the length of delay in diagnosis and the extent of damage incurred from severe acidosis and circulatory shock .


Because the disease is transmitted as an autosomal recessive trait, the male-to-female incidence is equivalent.


Clinical onset occurs shortly (within hours) after birth. Maternal ingestion of biotin supplements during pregnancy may alter the time of presentation by days or weeks.




As a water-soluble vitamin, biotin easily crosses the placenta; thus, the infant with holocarboxylase synthetase (HCS) deficiency in utero grows uneventfully.

Birth weight, Apgar scores, and initial physical examination findings are entirely normal.

Because of the autosomal recessive inheritance, a family history of the disorder is unlikely.[10]


The major clinical findings of holocarboxylase synthetase deficiency include severe ketoacidosis, exfoliative dermatitis, and hypoglycemia.

As in individuals with other organic acidemias, whether fed or fasted, the increased accumulation of ketoacids causes tachypnea, irritability, lethargy, vomiting, and, eventually, coma.

Exfoliative dermatitis is usually absent in the earliest stages of clinical presentation.

Observe for unusual odors in the urine.


Presentation must be distinguished from all other causes, such as other organic acidemias, sepsis, and galactosemia.

The gene locus has been mapped to band 21q22.1, with a limited number of allelic variants described. A particular allelic variant, the p.L216R mutation, results in an unstable protein with increased turnover and a clinical picture of relative biotin-resistance.[11] Treatment of these patients with massive amounts of daily biotin (>1 g) has resulted in relatively good outcomes.[12, 13]





Laboratory Studies

Initial studies in holocarboxylase synthetase (HCS) deficiency should include the following:

  • Serum electrolyte levels

  • Blood ammonia levels

  • Urinalysis

  • Culture samples appropriate for an evaluation for Sepsis

Aside from demonstration of an anion gap, hyperammonemia, and systemic acidosis, no routine laboratory tests are of use; in this context, urinary odor, if present, may be of critical importance in making a rapid diagnosis.

Urinary organic acid evaluation demonstrates the presence of lactate, beta-hydroxypropionate, beta-methylcrotonate, and beta-hydroxyisovalerate, as well as numerous other quantitatively less apparent organic acids.

Making a definitive biochemical diagnosis depends on demonstration of impaired activity of pyruvate, propionyl-CoA, and beta-methylcrotonyl-CoA carboxylases in white cells or cultured fibroblasts.

Techniques for direct sequencing of holocarboxylase synthetase gene mutations are available and should be performed following definitive diagnosis.

Prenatal diagnosis using chorionic villus biopsy samples and direct sequencing of fetal DNA has been reported.[14]



Medical Care

Because of the acute and fulminant initial presentation of holocarboxylase synthetase (HCS) deficiency, treatment is almost always initiated in the hospital.

Once treatment has begun and the acute signs and symptoms have disappeared, the child may be safely discharged and observed on an outpatient basis.

A powdered form of biotin supplementation may be better digested than tablets for the treatment of neonatal holocarboxylase deficiency.[15]


Surgical Care

No surgical procedures are necessary.


The patient and parents should consult a biochemical geneticist.


No dietary restrictions are needed.


No restriction is necessary.



Essential coenzymes

Class Summary

Essential coenzymes are organic substances the body requires in small amounts for various metabolic processes. Biotin is a coenzyme for 4 carboxylases.

Biotin (Vitamin H)

Water-soluble B vitamin that is important in metabolism of energy derived from food. Daily administration is essential to maintain metabolic equilibrium.



Further Outpatient Care

With regular biotin administration, frequent follow-up visits to a specialist should not be required for patients with holocarboxylase synthetase (HCS) deficiency.

Growth and development should be closely observed to address residual morbidity from the initial insult.

Inpatient & Outpatient Medications

Biotin is the only medication necessary for treatment.


Prolonged failure to adhere to the daily biotin requirement results in an insidious onset of the underlying disease, with potentially life-threatening consequences.


Early and consistent treatment allows for an excellent prognosis.

Patient Education

Families must not become sanguine during biotin administration because affected infants appear so healthy.

Teenaged patients often rebel against taking daily medication, despite the fact that biotin is an odorless, tasteless capsule.