John A. Sturman

Development of mammalian sulfur metabolism: absence of cystathionase in human fetal tissues.

Extract: Cystathionase activity was absent from human fetal liver and brain as early as 6 weeks of gestation. Hepatic methionine-activating enzyme (26 ± 3 nmoles/mg protein/hr) and hepatic cystathioninesynthase (21 ± 4 nmoles/mg protein/hr) were present (cf. 86 ±16 and 98 ± 19 nmoles/mg protein/hr, respectively, in mature human liver). All three activities were absent from the placenta. Human fetal liver contained higher concentrations of cystathionine (14 ± 2

Absence of cystathionase in human fetal liver: is cystine essential?

mUmoles/100 g wet weight) than mature human liver (0) and human fetal brain (4.0 ± 0.6

Development of Methyltransferase Activities of Human Fetal Tissues

mUmoles/100 g wet weight). Methionine-activating enzyme of human fetal brain, but not liver, showed a tendency to increase with development (coefficient of correlation was 0.62; 0.01 < P < 0.05).35S-l-methionine injected into the umbilical vein of six human fetuses was incorporated into free methionine in liver and brain, but not into free cyst(e)ine, homocyst(e)ine, taurine or, except for the smallest fetus, cystathionine. 35S-l-cysteine similarly injected was incorporated into free cysteine in liver and brain to a greater extent than in plasma, whereas it was incorporated into cystine in plasma to a greater extent than in either liver or brain. Incorporation of 35S into cystathionine in liver was greater from 35S-l-cysteine than from 35S-l-methionine. Both 35S-l-methionine and 36S-l-cysteine were actively incorporated into tissue proteins: methionine > cysteine and liver > kidney > brain.Incorporation of 35S-l-methionine and 35S-l-cysteine incubated with minced liver from four human fetuses showed more active incorporation of methionine (11,836–15,045 dpm/mg protein) than cysteine (7,044–9,856 dpm/mg protein).Speculation: These studies suggest that cysteine is an essential amino acid in human fetuses and in infants for some time after birth, especially if they were born prematurely.

Biochemical Observations on So-called Hereditary Tyrosinemia

Cystathionase activity is not measurable in the livers of 24 human fetuses and 3 premature infants, and the concentration of cystathionine in the liver is higher than that of the brain. The placenta does not subserve the trans-sulfuration function. Cystine (or cysteine) thus may be an essential amino acid in the immature human.

Taurine in developing rat brain: Transfer of [35S]taurine to pups via the milk

Extract: N5-Methyltetrahydrofolate-homocysteine methyltransferase specific activity was higher in fetal (2nd trimester) human liver and kidney (4.70 ± 0.20 and 7.25 ± 0.23 nmol/mg protein/hr) than in mature human liver and kidney (1.30 ± 0.16 and 0.76 ± 0.18 nmol/mg protein/hr). During the same period, there was a significant correlation of decreasing specific activity of this enzyme in fetal brain with increasing crown-rump length (r = −0.72; P < 0.005), reaching the specific enzymatic activity of adult brain (1.37 ± 0.26 nmol/mg protein/hr).Betaine-homocysteine methyltransferase specific activity was lower in fetal liver and brain (1.82 ± 0.21 and 0.20 ± 0.05 nmol/mg protein/hr) than in mature liver and brain (7.78 ± 1.89 and 0.37 ± 0.07 nmol/mg protein/hr). During the same period, there was a significant correlation of increasing enzymatic activity in fetal kidney with increasing crown-rump length (r = 0.80; P < 0.005) toward the mean specific activity of mature kidney (22.6 ± 2.0 nmol/mg protein/hr).Serine tetrahydrofolate 5,10-hydroxymethyltransferase specific activity showed no significant difference between fetal and mature liver and kidney; however, the fetal brain showed a significant correlation of decreasing specific activity of this enzyme with increasing crown-rump length (r = −0.69; P < 0.005).The specific activities of betaine-homocysteine methyltransferase and serine-tetra-hydrofolate 5,10-hydroxymethyltransferase in the liver of the neonate was not different from that in the mature liver. N5-Methyltetrahydrofolate-homocysteine methyltransferase in neonatal liver attained a specific activity similar to that found in mature liver before cystathionase did.Cystathionase in 2nd trimester human fetal kidney, in contrast to cystathionase in human fetal liver and brain, already has attained two-thirds of the mean specific activity of mature kidney.Speculation: Although the measurements of these enzyme activities as assayed in vitro do not take into consideration substrate availability in vivo, the relative specific activities of these methyltransferases in human fetal liver and brain, considered together with the absence of cystathionase, suggests to us that perhaps, in these two organs, the trans-sulfuration pathway for the further metabolism of homocysteine is turned off in favor of the N6-methyltetrahydrofolate-Bi2remethylation pathway (Fig. 1). The latter pathway converts 7V6-methyl tetrahydrofolate, the major monoglutamic folate in liver and serum [1], to tetrahydrofolate. The latter form of folate reacts with the f3 carbon of serine, on serine-tetrahydrofolate 5,10 hydroxymethyltransferase, to form Ni · 10-methylenetetrahydrofolate, a 1-carbon precursor for the de novo synthesis of thymidylate, which is uniquely required for DNA but not for RNA. This suggests that the /3 carbon of serine is being shunted into DNA synthesis during periods of rapid cellular multiplication, rather than having the entire carbon skeleton accept the sulfur from homocysteine to form cysteine. The latter thus becomes an essential amino acid in human fetal liver and brain.

Effects of deficiency of vitamin B6 on transsulfuration

Extract: Decreased activities of methionine-activating enzyme (ATP: L-methionine-S-adenosyl-transferanse, E.c. 2.5.1.6) (23 and 18 versus normal 86 nmoles product produced/mg soluble protein/h) and cystathionine synthetase [30] (28 and 6 versus normal of 98 nmoles product produced/mg soluble protein/h) in the presence of normal activity of cystathionase [30] (104 and 108 versus normal of 125 nmoles product produced/mg soluble protein/h) were demonstrated in the liver of two patients with so-called hereditary tyrosinemia. This decreased activity also was associated with documented deficiency of p-hydroxyphenylpyruvic acid oxidase, tyrosine transaminase, and phenylalanine hydroxylase with values of 3 and 1.2 versus normal of 60, 2.6 and 0.8 versus normal of 20, and 3.5 versus normal of 13.7 μmoles/g wet weight of liver/h, respectively (table II). In one patient, the biopsy was performed after the concentration of tyrosine in the plasma had been made normal (less than 1.0 mg/100 ml) for 3 months by dietary restriction (fig. 2). This patient has subsequently maintained normal concentrations of amino acids in the plasma on an ad libitum diet for 18 months. These findings give evidence that the abnormalities on the pathway of metabolism of methionine are independent of the abnormality in the metabolism of tyrosine and that the latter may be self-limiting in some cases (table I).Speculation: These studies suggest that the hypertyrosinemia and the hypermethioninemia seen in so-called here-ditary tyrosinemia are each nonspecific manifestations of a phenotype as yet unidentified. Whether or not methionine accumulates in the plasma in the presence of these defects in the transsulfuration pathway is probably a function of an enlarged free amino acid pool in the liver when protein synthesis in that organ is reduced.

Polyamine biosynthesis in human fetal liver and brain.

The concentration of taurine in rat milk is very high for the first few days after birth and then falls rapidly. [35S]Taurine injected intraperitoneally into lactating dams after birth was transferred via the milk to the pups, and accumulated in the brains of the pups to a greater extext than in the livers of the pups. Maximal accumulation of [35S]taurine so transferred to the brain of the pups was reached by 5 days after birth, and remained constant for at least 10 days beyond this poiht. The specific radioactivity in the brain of the pups also reached a maximal value at 5 days after birth and thereafter declined because of the expanding pool of unlabeled taurine in brain. At 5 days after birth, each pup has received approximately 4 mumol taurine from the mother via the milk, and a minimum of 7% of the total taurine in brain at this time originated from the milk. Speculation Even in the rat, a species which can synthesize taurine very easily from cysteine and methionine precursors, a significant amount of performed taurine is transferred to the developing animal via the milk. We suggest that the human infant, who cannot synthesize adequate taurine from cysteine and methionine precursors (9, 10, 40), may be dependent on its diet as a taurine source. Human milk contains a high concentration of taurine, whereas synthetic formulas contain virtually none. Taurine may be an essential nutrient for the rapidly growing human infant (and may be for the adult human also) and perhaps should be included as a supplement in synthetic formulas.

Transfer of cyst(e)ine and methionine across the human placenta.

Abstract Transsulfuration processes have been studied in vitamin B 6 -deficient and control rats. After five weeks, the major transsulfuration step affected is the cleavage of cystathionine, and this is due to decreased formation of holocystathionase and not of apocystathionase. In vitamin B 6 deficiency, cystathionine accumulates in plasma and urine, and in all of the tissues studied, as expected of a compound proximal to an enzymatic block. Taurine, a compound distal to the block, remains unchanged in all of the tissues studied and in plasma, but decreases in urine, establishing decreased renal clearance as the mechanism for conserving taurine.

Cysteine sulfinic acid decarboxylase in rat brain: Effect of vitamin B6 deficiency on soluble and particulate components

Extract: S-Adenosylmethionine decarboxylase specific activity in fetal (9.0–25.0 cm crown-rump length) human liver was 20-fold greater than in mature human liver, both in the absence of putrescine (63.1 ± 7.2 versus 3.5 ± 0.8 pmol CO2/mg protein/30 min) or in the presence of putrescine (116.9 ± 8.2 versus 6.2 ± 1.0 pmol CO2/mg protein/30 min). Extracts of fetal human brain did not have a significantly greater specific activity of S-adenosylmethionine decarboxylase than extracts of mature human brain when assayed in the absence of putrescine and had less when assayed in the presence of putrescine (84.7 ± 14.9 versus 175.5 ± 30.6 pmol CO2/mg protein/30 min). Ornithine decarboxylase specific activity was greater in fetal liver than in mature liver (8.8 ± 1.6 versus 1.1 ±0.3 pmol CO2/mg protein/30 min) and 20-fold greater in fetal brain than in mature brain (71.2 ± 12.1 versus 3.1 ± 1.4 pmol CO2/mg protein/30 min).The concentration of putrescine was threefold greater in fetal human liver than in mature human liver (21.6 ± 2.7 versus 6.5 ± 0.5 μmol/100 g) and eightfold greater in fetal human brain than in mature human brain (42.9 ± 2.3 versus 5.5 ± 0.6 μmol/100 g). The concentration of spermidine was fourfold greater in fetal human liver than in mature human liver (81.2 ±1.6 versus 21.4 ± 1.2 μmol/100 g), but its concentration in fetal human brain was not different from that in mature human brain. Spermine concentration was greater in fetal human liver than in mature human liver (107.3 ± 2.4 versus 70.7 ± 6.8 μmol/100 g) and greater in fetal human brain than in mature human brain (30.7 ±1.8 versus 22.1 ±0.9 μmol/100 g).Speculation: The pathway of methionine metabolism in human fetal liver and brain is adapted to conserve the sulfur of homocysteine by delayed development of the trans-sulfuration of homocysteine to cysteine. This adaptation may implement RNA synthesis, as well as protein synthesis and DNA synthesis.

Cystine metabolism in vitamin B6 deficiency: Evidence of muliple taurine pools☆

Extract: After intravenous loads in pregnant women, L-methionine, L-leucine, and L-ornithine were transferred from maternal to fetal plasma against a two- to threefold difference in initial concentration. Cyst(e)ine is unique among the free amino acids of plasma in that its basal concentration in maternal plasma was equal to or greater than that in fetal plasma. Furthermore, after intravenous loads with L-cystine or L-cysteine, total cyst(e)ine (cystine and cysteine) was transferred less readily to the fetal plasma. Although the concentrations of cystine in the fetal plasma continued to rise in the face of rapidly falling concentrations of cystine in the maternal plasma, at no time during the experiment did the concentration of cystine in fetal plasma exceed that in the maternal plasma. When D-cystine was administered intravenously to a mother, in amounts equimolar with L-cystine, the transfer of D-cystine was not measurable.Speculation: Cyst(e)ine is the end-product of the transsulfuration pathway, which transfers the sulfur from the 4-carbon skeleton of methionine to the 3-carbon skeleton of serine. Methionine-actvating enzyme, the first enzyme on this pathway, is inhibited by cyst(e)ine [2, 5, 20]. It is also allosteric [19], a characteristic often associated with negative feedback inhibition. Cystathionase, which cleaves cystathionine to cysteine and α-ketobutyrate and is the last enzyme on this pathway, is absent from the liver and brain of the human fetus. Therefore, we postulate that the special mechanism for the transfer of cyst(e)ine across the human placenta is an adaptation to control the transfer of sufficient maternal cyst(e)ine for synthesis of protein and glutathionine without undue inhibition of the activation of methionine to S-adenosylmethionine, a compound involved in important synthetic reactions.