Cystathionine cleavage enzyme

Binkley and Okeson purified the enzyme system that cleaves cystathionine and found that neither phosphate nor ATP was required for activity, thus correcting the previous report that ATP was required. In addition to splitting cystathionine, this enzyme preparation also produced H2S from cysteine. The authors suggest that their enzyme may be identical with cysteine desulfhydrase. Binkley also reported that he had been able to synthesize cystathionine enzymatically from homocysteine and serine by a fractionated liver preparation which had been freed from the cystathionine cleavage enzyme, serine dehydrase and homoserine deaminase. The activity of the enzyme synthesizing cystathionine was either inhibited or unaffected by ATP, DPN, AMP, and various metal ions. 

The formation of a-ketobutyric acid from homoserine (reaction 5) is based on the observation of Carroll et al. that homoserine or its lactone could be converted to this keto acid by the action of liver extracts under anaerobic conditions, whereas a-aminobutyric acid did not produce any keto acid upon incubation with the enzyme preparation under the conditions of cystathionine cleavage. 

Be deficiency was found to reduce transfer of sulfur from homocysteine to serine, and this could be restored in liver extracts by the addition of pyridoxal phosphate. Binkley et oZ. reported the activation of both the cystathionine-synthesizing enzyme and the cleavage enzymes by pyridoxal phosphate. 

Cleavage of cystathionine is accomplished by cystathi-onase, another pyridoxine-dependent enzyme that is coded on human chromosome 16 (Fig. 40-4 reaction 6). The enzyme functions almost entirely to produce cysteine, there being virtually no reversal of the reaction. 

The subsequent cleavage of cystathionine to yield cysteine, a-ketobutyrate and NH4+ is catalyzed by y-cystathionase, a pyridoxal-phosphate-containing enzyme. This transsulfura-tion pathway is one of the routes used for methionine catabolism. 

Transamination is just one of a wide range of amino acid transformations that are catalyzed by PLP enzymes. The other reactions catalyzed by PLP enzymes at the a-carbon atom of amino acids are decarboxylations, deaminations, racemizations, and aldol cleavages (Figure 23.13). In addition, PLP enzymes catalyze elimination and replacement reactions at the p-carbon atom (e.g., tryptophan synthetase p. 696) and the y-carbon atom (e.g., cystathionine p-synthase, p. 693) of amino acid substrates. Three common features of PLl catalysis underlie these diverse reactions. 

Autotrophic organisms synthesize methionine from asparfafe as shovm in the lower right side of Fig. 24-13. This involves fransfer of a sulfur atom from cysfeine info homocysfeine, using the carbon skeleton of homoserine, the intermediate cystathionine, and two PLP-dependent enzymes, cystathionine y-synthase and cystathionine p-lyase. This transsulfuration sequence (Fig. 24-13, Eq. 14-33) is essentially irreversible because of the cleavage to pyruvate and NH4+ by the P-lyase. Nevertheless, this transsulfuration pathway operates in reverse in the animal body, which uses two different PLP enzymes, cystathionine P s3mthase (which also contains a bound heme) and cystathionine y-lyase (Figs. 24-13,24-16, steps h and i), in a pathway that metabolizes excess methionine. 

As noted above, cystathionine formation is the other major fate of methionine. The condensation of homocysteine with serine is catalyzed by the vitamin requiring enzyme cystathionine P-synthase. In the last step of the transsulfuration sequence, cystathionine undergoes cleavage to cysteine and a-ketobutyrate in yet another enzyme reaction that requires pyridoxal phosphate. 

In addition to cysteine, it has been observed that a-ketobutyric acid is a product of the enzymatic cleavage of cystathionine. This could have been formed from homoserine, since the latter is converted to an a-keto acid by liver extract under anaerobic conditions. An enzyme that specifically deaminates homoserine has been reported to be present in liver by Binkley and Olson. ... 

A yellow enzyme has been isolated from rat liver in crystalline form which deaminates homoserine giving rise to a-ketobutyrate and ammonia (235). The same preparation catalyzes the cleavage of cystathionine to cysteine. The probable identity of homoserine deaminase and cystathi-onase is proposed. The crystalline enzyme contains pyridoxal phosphate as the prosthetic group. [One mole of enzyme contains 4 moles of pyridoxal phosphate ( 3 ).] No significant quantity of bound heavy metals ions could be detected. The resolution of the enzyme into pyridoxal phosphate and oatalytically inactive apoenzyme and subsequent reconstitution of the active enzyme by combining the two components 2S6a) provides conclusive... 

Rhizobitoxine, an analogue of cystathionine produced by certain strains of Rhizobium japonicum (Owens et al., 1972), is a potent irreversible inhibitor of plant cystathionine 8-lyase both in vivo (Giovanelli et al., 1973) and in vitro. Inhibition of the purified spinach enzyme was of the active-site-directed irreversible type (Giovanelli et al., 1971), and probably involves covalent linkage of a cleavage product of rhizobitoxine to the pyridoxal phosphate prosthetic group of the enzyme. 

The major developmental change which takes place In both brain and liver is the postnatal activation of the transsulfuration pathway of methionine metabolism. The net result of this pathway is the transfer of the sulfur atom from homocysteine to the carbon skeleton of serine to form cysteine. This conversion is mediated by two enzymes cystathionine synthase (L-serine hydro-lyase adding homocysteine, EC 4.2.1.22) which catalyzes the 3-activation of serine and the addition of homocysteine to form the thio-ether, cystathionine cystathionase (EC 4.4.1.1) which catalyzes the y-cleavage of cystathionine to form cysteine (Fig. 1). Both of these enzymes catalyze reactions other than those described above although their importance vivo is uncertain (Tallan et al., 1974). In mature mammals, activities both of cystathionine synthase and of cystathionase are present in brain and liver, although cystathionase activity in... 

Their purified enzyme also supported the cleavage of cystathionine according to the following reaction ... 

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