In a cystine-free medium supplemented with cystathionine (0.5 mM), three longterm lymphoid (LTL) lines from normal subjects incorporated [14C]-leucine into protein almost as well as in cystine-containing control medium. However, lines from 2 patients with B6-responsive cystathioninuria and a line from 1 patient with B(6)-unresponsive cystathioninuria failed to incorporate [14C]-leucine under these conditions. Addition of pyridoxine up to 0.5 mM had no consistent effect on cells from normal subjects. Addition of 0.5 mM pyridoxine to the cystine-free medium containing cystathionine afforded leucine uptake up to 80% of the control with one B6-responsive line and 20% with the other. The B6-unresponsive line failed to incorporate leucine even in the pyridoxine-enriched medium. Mehionine, a key amino acid involved in many fundamental processes in mammalian metabolism is converted to cyst(e)ine via the transsulfuration pathway [4]. The last step on this pathway is catalyzed by y-cystathionase (L-cystathionine cysteine-lyase [deaminating],EC 4.4.1.1), for which the cofactor is pyridoxal phosphate (PLP), a coenzymatically active form of B(6) [8]. Primary cystathioninuria is an inborn error of sulfur metabolism resulting from an inherited deficiency of cystathionase. The condition was first reported in a patient with severe mental deficiency [5]. Numerous other cases with a variety of clinical symptoms have been reported [12, 13], This lack of a distinct clinical picture has led to the suggestion that mental deficiency and other clinical manifestations are not causally related to the enzymatic defect. Only 2 of these cases have not been B(6)-responsive [7, 15], Thus, the relation of clinical manifestations to enzymatic deficiency in the cases unresponsive to B(6) remains a question. Utilizing immunochemical methods, Pascal et al. [9, 10] have shown that different mutations affect the cystathionase molecule. One form of cystathioninuria, B(6)-unresponsive,appears to result from the absence of synthesis of the enzyme protein. B(6)-responsive forms of cystathioninuria result from production of cystathionase molecules having altered ability to combine with coenzyme but having antigenic identity with the enzyme as shown by agar double diffusion analysis. In some cases there may be changes in antibody-binding capacity as well. Using a special stain for cystathionase activity, the coenzyme, PLP, was shown to act directly on the cystathionase molecule [11]. The studies described in this paper were performed in order to define requirements for cultivation of LTL cells in a cystine-free medium, using cells from normal subjects and from patients with cystathioninuria, and to develop a system for studying the effects of cystathionase deficiency in a living system. For this purpose LTL cells are more suitable than skin fibroblasts since cystathionase activity is considerably less in fibroblasts [9]. We demonstrate that LTL lines are a useful model for such studies. Other investigators [6] have used LTL lines to study cystathionase in cells from leukemic patients.

Keywords:
Cystathionase, Cystathioninuria, Enzyme structrally altered, Coenzyme
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1982
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