Plasma methionine

Figure 32, Chromatograms of plasma and urine samples with various abnormalities, A, Phenylalaninemia B, tyrosinemia C, elevated plasma methionine seen in homocystinuria D, glycinemia E, normal urine F, argininosuccinic aciduria G, homocystinuria H, hyperglycinuria I, hyperlysinuria.

Fukagawa, N., Ajami, A-, and Young, V. R. (1996). Plasma methionine and cysteine kinetics in response to an intravenous glutathione infusion in adult humans. Am.. Physiol. 270, E209-E214. 

Deficiencies of methionine adenosyltransferase, cystathionine 8-synthase, and cystathionine )/-lyase have been described. The first leads to hypermethioninemia but no other clinical abnormality. The second leads to hypermethioninemia, hyperhomocysteinemia, and homo-cystinuria. The disorder is transmitted as an autosomal recessive trait. Its clinical manifestations may include skeletal abnormalities, mental retardation, ectopia lentis (lens dislocation), malar flush, and susceptibility to arterial and venous thromboembolism. Some patients show reduction in plasma methionine and homocysteine concentrations and in urinary homocysteine excretion after large doses of pyridoxine. Homocystinuria can also result from a deficiency of cobalamin (vitamin B12) or folate metabolism. The third, an autosomal recessive trait, leads to cystathioninuria and no other characteristic clinical abnormality. 

A nutritional molybdenum deficiency with clinical symptoms similar to those of sulfite oxidase deficiency was identified in a human patient receiving long-term total parenteral nutrition (TPN) (Abumurad et al. 1981). The clinical symptoms included irritability leading to coma, tachycardia, tachypnea, and night blindness. A reduced intake of protein and sulfur-amino acids alleviated the symptoms, but they were aggravated by infusion of sulfite. The biochemical findings were low tissue sulfite oxidase activity a 25-fold increase in thiosulfate excretion a 70% reduction in urinary output of sulfate and a marked rise in plasma methionine. The clinical symptoms of molybdenum deficiency were totally eliminated by daily supplementation of 300 pg of the element. 

Innis, S.M. A. Davidson F. George S. Melynk S.J. James. Choline-related supplements improve abnormal plasma methionine- homocysteine metabolites and glutathione status in children with cystic fibrosis. Am. J. Clin. Nutr. 2007, 85, 702—708. 

As the name implies, renal clearance of abnormal levels of homocystine in the plasma causes excessive excretion of the amino acid in the urine. In cystathionine P-synthase deficiency, plasma methionine concentrations are elevated as well -this serves as a point of distinction from the remethylation defects. At present, it appears that the pyridoxal phosphate response may be explained by the fact that this vitamin increases the steady-state concentration of the active enzymes by decreasing the rate of apoenzyme degradation and possibly by increasing the rate of holoenzyme formation. The explanation is not entirely satisfactory, however, since in vitro studies have shown detectable levels of enzyme activity in mutant fibroblasts that have no response, while in other mutant lines without detectable enzyme activity, response has occurred. Once again, a distressing lack of correspondence between in vivo observations and in vitro experiments forces investigators to probe the secrets of these diseases more deeply. 

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. 

Fibroblasts from each patient had markedly reduced levels of the above enzyme. The defect results in an inability to synthesize 5-methyltetrahydrofolate in amounts sufficient for the remethylation of homocystine to methionine. Homocystine accumulates in plasma, and the plasma methionine is decreased. As in the other remethylation defect, there is accumulation of cystathionine. Treatment with high doses of folic acid has been beneficial in several patients (Mudd et al., 1989). 

The diagnosis of homocystinuria is based on the recognition of the clinical phenotype in conjunction with the identification of an elevated total plasma homocysteine and elevated plasma methionine concentrations (via quantitative plasma amino acid analysis). Low cystine and low cystathionine are also seen (Box 14.3). In addition, increased urinary excretion of homocysteine as well as cysteine-homocysteine disulfide can be identified on urine amino acid analysis. Confirmation of the diagnosis can be done via enzyme assay, typically performed on cultured skin fibroblasts, lymphocytes, or liver tissue, or via molecular studies. 

A number of conditions outside the scope of this chapter are associated with mild hyperhomocysteinemia, including premature vascular disease and deficiencies of folate and vitamin B12 [7]. Oral methionine loading (0.1 g/kg) may be used to investigate such patients. Postloading plasma homocysteine levels reach a peak at 4-8 h and approach preloading values within 2-4 days. Together with plasma methionine, the baseline level and postloading rise of plasma homocysteine may provide information on the various inherited and acquired defects of homocysteine metabolism. 

Table 2. Blood plasma methionine (Met, mg/100 ml), lysine (Lys, mg/100 ml) and urea (nmol/l) the day before (day 0) and 5 days after start of feeding ruminal protected D,L-Met and L-Lys HCl products (Timet and Relys , respectively).

Homocysteine arises from dietary methionine. High levels of homocysteiae (hyperhomocysteinemia) are a risk factor for occlusive vascular diseases including atherosclerosis and thrombosis (81—84). In a controlled study, semm folate concentrations of <9.2 nmol/L were linked with elevated levels of plasma homocysteiae. Elevated homocysteine levels have beea associated also with ischemic stroke (9). The mechanism by which high levels of homocysteine produce vascular damage are, as of yet, aot completely uaderstood. lateractioa of homocysteiae with platelets or eadothehal cells has beea proposed as a possible mechanism. Clinically, homocysteine levels can be lowered by administration of vitamin B, vitamin B 2> foHc acid. 

Methyl-tetrahydro folic acid is furthermore, together with vitamin B12 and B6, required to regenerate homocysteine (see Vitamin B12, Fig. 1). Homocysteine results when methionine is used as a substrate for methyl group transfer. During the last few years, homocysteine has been acknowledged as an independent risk factor in atherosclerosis etiology. Folic acid supplementation can help reduce elevated homocysteine plasma levels and is therefore supposed to reduce the risk of atherosclerosis as well [2]. 

Contrary to LDL, high-density lipoproteins (HDL) prevent atherosclerosis, and therefore, their plasma levels inversely correlate with the risk of developing coronary artery disease. HDL antiatherogenic activity is apparently due to the removal of cholesterol from peripheral tissues and its transport to the liver for excretion. In addition, HDL acts as antioxidants, inhibiting copper- or endothelial cell-induced LDL oxidation [180], It was found that HDL lipids are oxidized easier than LDL lipids by peroxyl radicals [181]. HDL also protects LDL by the reduction of cholesteryl ester hydroperoxides to corresponding hydroperoxides. During this process, HDL specific methionine residues in apolipoproteins AI and All are oxidized [182]. 

Homocystinuria can be treated in some cases by the administration of pyridoxine (vitamin Bs), which is a cofactor for the cystathionine synthase reaction. Some patients respond to the administration of pharmacological doses of pyridoxine (25-100 mg daily) with a reduction of plasma homocysteine and methionine. Pyridoxine responsiveness appears to be hereditary, with sibs tending to show a concordant pattern and a milder clinical syndrome. Pyridoxine sensitivity can be documented by enzyme assay in skin fibroblasts. The precise biochemical mechanism of the pyridoxine effect is not well understood but it may not reflect a mutation resulting in diminished affinity of the enzyme for cofactor, because even high concentrations of pyridoxal phosphate do not restore mutant enzyme activity to a control level. 

The Ras proteins are synthesized as biologically inactive, cytosolic precursor proteins. They are then modified by several post-translational processing steps at the carboxyl terminal end and thereby converted into biologically active proteins localized at the plasma membrane. The cysteine of the C-terminal CAAX sequence (C is cysteine, A is generally an aliphatic amino acid, and X is methionine, serine, alanine, or glutamine) is first enzymatically S-farnesylated the AAX part is then cleaved off by a specific protease, and the free C-terminal cysteine is finally converted into a methyl ester (Scheme 1). 

The central unit of these peptidomimetics imitates a /1-turn and brings the NH2-terminus of the cysteine analogue and the CO OH terminus of the methionine in spatial proximity these can then complex the Zn2+ ion which is essential for activity of the FTase [26]. The free acid 7 inhibits the enzyme with an IC50 value of 1 nmol/1, whilst in intact cells the methyl ester 8, despite its weaker in vitro activity, is significantly more potent because it can penetrate the plasma membrane better due to its lower polarity. This property can be used to convert the morphology of H-Ras-transformed cells back to the normal form and to inhibit growth of these cells, whereas the substance shows no effect on Src-transformed and untransformed rat fibroblasts. The inhibitor therefore acts selectively on transformed cells and does not influence growth of normal cells. This result is noteworthy because farnesylation of the wild type H-Ras protein... 

In rat blood and in the cell culture medium RPMI 1640 (+15% fetal calf serum) oxaliplatin (4) forms the major biotransformation products [Pt(dach)Cl2], [Pt(dach)(H20)Cl]+ (only in plasma ultrafiltrate), and [Pt(dach)(methionine)]+ (91). 

A T-C polymorphism was described in the sequence of apo(a)-KIV10 (V6), corresponding to nucleotide 12,605 of the published cDNA sequence (M24). This variant results in the substitution of a methionine (ATG) with a threonine (ACG) at this position. No correlation was observed between the polymorphism and plasma Lp(a) levels. Although the Met-Thr substitution is present within the lysine binding pocket in KIV10, its effect on lysine binding properties of this kringle remains to be determined. 

A study involving twenty-six laboratories was carried out to assess the quality of amino acid analysis, using samples of urine and lyophilized plasma. Coefficients of variation ranged from 13% for glycine to 65% for methionine. Automated IEC followed by ninhy-drin detection (37) seemed to perform better than other methods however, there was no clearly superior method and no analyzer clearly outperformed the others. This seems to point to the importance of personal proficiency and expertise in the performance of such analyses137. 

Determination of iodo amino acids by HPLC with inductively coupled plasma (ICP)-MS detection had LOD 35-130 pg of I, which is about one order of magnitude lower than with UVD usually applied for these compounds175. Amino acids and peptides containing sulfur, such as cysteine, cystine, methionine and glutathione, can be determined after HPLC separation by pulsed electrochemical detection, using gold electrodes176. 

Husek P, Matucha P, Vrankova A, Simek P. 2003. Simple plasma work-up for a fast chromatographic analysis of homocysteine, cysteine, methionine and aromatic amino acids. J Chromatogr B 789 311. 

Answer C Only methionine is degraded via the homocysteine/cystathionine pathway and would be elevated in the plasma of a cystathionine synthase-deficient patient via activation of homocysteine methyl-transferase by excess substrate. 

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