Evidence that 4-aminobutyric acid and 5-aminolevulinic acid share a common transport system into Saccharomyces cerevisiae.

It has been previously reported that 5-aminolevulinic acid (ALA) and 4-aminobutyric acid (GABA) share a common permease in Saccharomyces cerevisiae (Bermúdez Moretti et al., 1993). The aim of the present work was to determine the relationship between the transport of these compounds in isolated cells. Assessment of amino acid incorporation was performed in S. cerevisiae using 14C-ALA or 3H-GABA. Initial rates of ALA incorporation in cells grown in the presence of 5 mM ALA and 5 mM GABA, were three to four times lower than in cells grown without supplements. Kinetic studies indicate that GABA competitively inhibits ALA transport. During the growth phase GABA uptake was also inhibited by 74% and 60% in the presence of ALA and GABA, respectively. These findings indicate that in S. cerevisiae the structurally related compounds, ALA and GABA, may be incorporated into the cells by a common carrier protein. Should this occur in other lukaryotic cells it may explain the neurotoxic effect attributed to ALA in the pathogenesis of acute porphyrias.

Studies on porphobilinogen-deaminase from Saccharomyces cerevisiae.

Porphobilinogen-deaminase from Saccharomyces cerevisiae has been isolated and partially purified 80- and 230-fold in the absence or presence of phenylmethylsulphonyl fluoride, respectively. Some properties of the isolated enzyme were studied. Porphyrin formation was linear with time and protein concentration. Optimum pH was about 7.5-7.8. Molecular mass of the protein was 30,000 +/- 3000 Dalton when the enzyme was purified in the presence of phenylmethylsulphonyl fluoride. A less active and unstable 20,000 Da molecular mass species was obtained when purification was performed in the absence of the protease inhibitor. Porphobilinogen-deaminase exhibited classical Michaelis-Menten kinetics. The apparent Km for uroporphyrinogen formation was 19 microM; Vmax was 3.6 nmol uroporphyrin/h and the Hill coefficient was n = 1. Also the action of several reagents on the activity was studied. Protective thiol agents had no effect. Heavy metals inhibited both porphyrin formation and porphobilinogen consumption, but known sulphydryl inactivating chemicals inhibit the former without modifying the latter. Ammonium ions had no effect on the activity while hydroxylamine completely inhibited both porphyrin formation and porphobilinogen consumption.

Different porphobilinogen-deaminase forms in wild and mutant strains of Saccharomyces cerevisiae. A possible correlation with its segregants behaviour.

1. Different porphobilinogen-deaminase (PBG-D) enzyme forms were found for D 27 and D 27/C6 (HEM R+) strains of Saccharomyces cerevisiae. 2. PBG-D was partially purified and chromatographed on Sephadex G-100 in either the presence or absence of a protease inhibitor. For D 27 only one active peak was observed while for D 27/C6 strain two active peaks were found. 3. A correlation between this differential behaviour and the presence of HEM R+ gene was looked for employing two segregants of one tetrad from D 27 and D 27/C6 mating.