Homocystine, and homocysteine

Cystathionine fisynthase (EC 4.2.1.22). Failure to form cystathionine from homocysteine and serine. Elevated homocysteine and methionine in serum. Urine contains homocystine and homocysteine-cysteine disulfide. Cystathionine virtually absent from brain, where it is normally present in significant quantities. Mental retardation. Lens detachment. Skeletal abnormalities (tall stature, arachnodactyly). Arterial and venous thrombosis. 

Reaction 4 is catalysed by cystathionine synthase (EC 4.2.1.13), an enzyme widely distributed in the tissues. In homocystinuria, cystathionine synthase is virtually completely absent or inactive in all tissues examined liver, brain and fibroblasts grown in tissue culture [33]. In some cases 1 to 2% of the normal enzymic activity can be demonstrated, in others no enzymic activity has been found [34]. As a result of the metabolic block, homocysteine accumulates and is partly converted to homocystine, partly to homocysteine-cysteine mixed disulphide and partly S-methylated to methionine by reactions 6 and 7 with, respectively, N -methyltetrahydrofolic acid and betaine as methyl donors. In infancy methionine and homocysteine are present in high concentrations in the plasma while homocystine and homocysteine-cysteine mixed disulphide are excreted in the urine later the concentration of methionine in the plasma drops. Cystathionine is normally present in highest concentration in the cells of the brain, though traces are found elsewhere and in the urine in homocystinuria no cystathionine can usually be demonstrated in the brain or urine [35]. The body s cysteine and cystine are also largely biosynthesized from methionine, though some is obtained from cysteine and cystine in dietary proteins in homocystinuria, cysteine/cystine becomes an essential amino acid. 

Cystathionine synthase has pyridoxal phosphate as cofactor a radically different form of treatment was introduced in 1967 giving pyridoxine at a dosage level of 50 to 200 mg per day [40]. The concentrations of methionine and homocysteine in the blood, and of homocystine and homocysteine-cysteine mixed disulphide in the urine, fell sharply on such treatment in... 

Rabenstein and Yamashita [52] determined penicillamine and its symmetrical and mixed disulfides by HPLC in biological fluids. Plasma and urine were deproteinized with trichloroacetic acid, and HPLC was performed on a column (25 cm x 4.6 mm) or Biophase ODS (5 pm) with a mobile phase comprising 0.1 M phosphate buffer (pH 3) and 0.34 mM Na octylsulfate at 1 mL/min. Detection was with a dual Hg-Au amalgam electrode versus a Ag-AgCl reference electrode. (z>)-penicillamine and homocysteine were determined at the downstream electrode at +0.15 V, and homocystine, penicillamine-homocysteine, and penicillamine disulfides were first reduced... 

In homocystinuria, cystathionine synthase is defective. Therefore, homocysteine does not react with serine to form cysteine (see Figure 7-10). The homocysteine that accumulates is oxidized to homocystine and excreted in the urine. Some cases respond to increased doses of vitamin B6, which forms pyridoxal phosphate, the cofactor for the synthase enzyme. 

Cystathionine synthetase, a pyridoxal phosphate (vitamin Be) enzyme, catalyzes the condensation of serine and homocysteine to form cystathionine. A deficiency of this enzyme leads to a buildup of homocysteine, which oxidizes to form homocystine. This may result in mental retardation, but sometimes causes dislocated lenses and a tall, asthenic build reminiscent of Marfan s syndrome. Patients with homocystinuria also have a clotting diathesis, requiring care to avoid dehydration during anesthesia. Their cysteine deficiency must be made up from dietary sources. In some cases, dietary intake... 

If the blood levels of methionine and homocysteine are very elevated and cystine is low, cystathionine p-synthase could be defective, but a cystathionase deficiency is also a possibility. With a deficiency of either of these enzymes, cysteine could not be synthesized, and levels of homocysteine would rise. Homocysteine would be converted to methionine by reactions that require B12 and tetrahydrofolate (see Chapter 40). In addition, it would be oxidized to homocystine, which would appear in the urine. The levels of cysteine (measured as its oxidation product cystine) would be low. A measurement of serum cystathionine levels would help to distinguish between a cystathionase or cystathionine p-synthase deficiency. 

Kleinman and Richie used the in-series approach with gold amalgam electrodes to measure 12 thiols and disulfides, including GSH, GSSG, cysteine, cystine, homocysteine, homocystine and several mixed disulfides in rat tissues. The first (upstream) electrode was maintained at —1.0 V and the second (downstream)... 

A similar separation of 50 physiological amino adds can be obtained with the highly efficient 3 pm cation exchanger from Hitachi (Tokyo, Japan) in about 2 h. The Hitachi column is characterized by an extremely low back pressure of 370-800 psi over the course of the run. In addition to key amino adds related to certain metabolic errors, less common amino acids such as homocysteine, homocitrulline, //o-isoleucine, argininosuccinic acid, cysteine-homocysteine mixed disulfides, homocystine, and argininosuccinic acid anhydride can also be separated under similar chromatographic conditions. 

The complete system contained per milliliter of solution 30 micromoles of dibasic ammonium phosphate, 0.2 micromole of DPN, 40 y of GSSG, 100 micromoles of alcohol, 200 y of alcohol dehydrogenase, 54 y of GSSG reductase, 0.7 micromole of homocystine, and 1.1 mg. of partially purified transhydrogenase.. fter 30 min. at 37° the mixture was analyzed for homocysteine. 

These include the natural amino acids, cystine and methionine, and their derivatives, cysteine, homocysteine, homocystine, and the tripeptide glutathione. 

When administered to subjects of the metabolic disease cystinuria, cysteine, homocysteine and methionine are excreted largely as additional cystine whereas administered cystine, homocystine and glutathione are almost completely oxidised. From these observations it is concluded that cystine can be metabolised without previous reduction to cysteine, and that glutathione can be meta-bobsed without previous hydrolysis, indicating that the metabolic history of an amino acid may depend on whether it is free or combined. Methionine, previous to its conversion into cysteine is demethylated to form homocysteine, which may undergo condensation to homocystine or degradation to simpler products. 

A. DIRECT OXIDATION OF THE SULFUR OF HOMOCYSTEINE 1. Oxidation of Homocysteine to Homocystine and the Fate of Homocystine... 

The biosynthesis of adenosine is theoretically controlled by several processes namely (1) the biosynthesis of adenosine from AMP by 5 -nucleotidase [EC 3.1.3.5], (2) from S-adenosyl homocysteine by S-adenosyl homocystine hydrolase [EC 3.3.1.1], (3) the metabolism of adenosine to AMP by adenosine kinase [EC 2.7.1.20], and (4) to inosine by adenosine deaminase (ADA) [EC 3.5.4.2], Interestingly, both 5 -nucleotidase and ADA activities were found to be highest in the leptomeninges of rat brain in contrast, the adenosine kinase activity was widely distributed throughout the brain parenchyma, which has negligible ADA activity... 

Homocysteine is a nonprotein-building amino acid formed as a metabolite in the methionine cycle. It was first associated with disease in 1962 (1,2). Individuals with a mutation in cystathionine-(3-synthase (CBS) develop classical homocystin-uria with extremely elevated plasma tHcy (> 100 xmol/L) (3). Homocystinuria is characterized by early atherosclerosis and thromboembolism as well as mental retardation and osteoporosis and is ameliorated by vitamin supplementation aimed at reducing the blood concentration of homocysteine (4). 

There are two pyridoxal phosphate-requiring enzymes in the homocysteine degradation pathway, which are associated with genetic diseases. In homo-cystinuria, cystathionine synthase is defective, and large amounts of homocystine are excreted in the urine. Some homocystinurics respond to the administration of large doses of vitamin B6. In cystathioninuria, cystathionase is either defective or absent. These patients excrete cystathionine in the urine. Cystathionase is often underactive in the newborns with immature livers, and cysteine and cystine become essential amino acids. Human milk protein is especially rich in cysteine, presumably to prepare the newborn for such a contingency. 

The fraction of intracellular homocysteine that does not undergo transsulfuration or remethylation is secreted into the extracellular space and ultimately finds its way into the blood. One major source of blood homocysteine is the liver, but some homocysteine is secreted into the blood by endothelial cells, circulating blood cells, and other tissues. Only about 2% of homocysteine in blood remains in its reduced, thiol form. The remainder circulates as a variety of different oxidation adducts, which include the disulfide, homocystine, as well as homocysteine-cysteine mixed disulfide and several protein-bound disulfides.About 70% of total homocysteine in blood is bound to the protein albumin through a disulfide linkage.When blood homocysteine is measured in the clinical laboratory, a reducing agent is added to the sample... 

Homocystine may induce Ap accumulation, synaptic dysfunction, and memory impairment. Lu et al. [581] reported that Hydroxysafflor Yellow A (HSYA) attenuates Ap accumulation, improves synaptic function, and reverses homocysteine-induced cognitive impairment. 

In plasma, homocysteine is present as both free (< 1 %) and oxidized forms (>99%). The oxidized forms include protein (primarily albumin)-bound homocysteine mixed disulfide (80-90%), homocysteine-cysteine mixed disulfide (5-10%), and homocystine (5-10%). Several studies have shown the relationship between homocysteine and altered endothelial cell function leading to thrombosis. Thus, hyperhomocysteinemia appears to be an independent risk factor for occlusive vascular disease. Five to ten percent of the general population have mild hyperhomocysteinemia. 

Transmethylation yields 1 mole of adenosine for every mole of methylated product. Patients are known with inborn errors that lead to incomplete metabolism of the homocysteine released after transmethylation. Homocysteinurics release about 3 mg of homocysteine per kilogram on unsupplemented diets, more on supplemented (G3, LI). This is minimal, owing to other fates of homocysteine and possible incompleteness of the block (L2). Cystathioninurics have been reported to excrete over a gram of cystathionine per day and up to 0.6 g per gram of creatinine (F6, H3, K9). Since the latter probably have some cystadiioninase, and some remethylation of homocystine may occur, the reported values are also minimal. Nevertheless, the moles of homocysteine and its products are of the same order as the amount of hypoxanthine formed per day. 

Methionine is not completely stable towards strong acid. A loss of 1274% by refluxing with 20% HCl for 20 h has been reported (Osono, 1955). The first decomposition product is homocysteine or homocystine (Osono et al., 1955). Homocysteic acid, aspartic acid, methionine sulfoxide, and methionine sulfone have also been identified as decomposition products (Pikkarainen and Kulonen, 1969) but oxidative processes were involved in these transformations. 

Elevated homocysteine (Hey) blood levels but also the presence of Homocystine in urine is the biochemical hallmark of these disorders and can be detected by a positive urinary cyanide nitroprnsside reaction. As other disulfides, including cystine and P-mercaptolactate cystine, also react, amino acid paper thin layer chromatography will be reqnired to distingnish these componnds from homocystine. Since there are two other forms of homocystinuria (disenssed later) in which plasma methionine is decreased, plasma amino acid evalnation is also in order. 

Patients have been described with this defect in remethylation of homocysteine. The initial patient presented a catastrophically ill newborn who died when 7 weeks old. While homocystine levels were elevated in blood and urine, the level of methionine in blood was quite low. Of great interest was the presence of large amounts of methylmalonic add in urine. Three other patients presented later in childhood, two of whom were retarded mentally. One of these patients also had severe megaloblastic anemia (Doscherholmen and Hagen, 1957). 

Homocysteine is an intermediate sulfur amino acid. Normal fasting homocysteine levels in plasma are between 5 and 15 pmol/1. Hyperhomocysteinemia are considered to be moderate between 16 and 30 pmol/l, intermediate (31 lOOpmol/1) and severe > 100 pmol/1. In plasma, 70-80% of homocysteine is bound to plasma proteins and 20-30% is free homocysteine circulating as homocysteine disulfide (homocystine) or mixed disulfides (cysteine-homocysteine). Folate deficiency leads to the decrease of homocysteine... 

An inborn error of metabolism in which homocystine is excreted in the urine due to a deficiency of cystathionine synthase, the enzyme which catalyses the formation of cystathionine from homocysteine and serine. Homocysteine is a sulphur containing amino acid and can be detected in the urine by tests for this type of amino acid. Among the symptoms of the condition are minor congenital abnormalities, mental retardation and dislocated lenses. 

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