Transport and metabolism of N-δ-chloroacetyl- l-ornithine by Saccharomyces cerevisiae

Abstract

The general amino acid transport system of Saccharomyces cerevisiae functions in the uptake of neutral, basic, and acidic amino acids (M. Grenson, C. Hou, and M. Crabeel, 1970, J. Bacteriol. 103, 770–777; J. Rytka, 1975, J. Bacteriol. 121, 562–570; C. Darte and M. Grenson, 1975, Biochem. Biophys. Res. Commun. 67, 1028–1033). We have previously demonstrated that this transport system can be inhibited by the amino acid, N-δ-chloroacetyl- l-ornithine (NCAO) (F. S., Larimore and R.J. Roon, 1978, Biochemistry 17, 431–436). In the present study radiolabeled NCAO was synthesized and its transport and metabolism studied. Under initial rate conditions: (a) NCAO was transported by the general amino acid transport system with a K m of 52 μ m, a V of 32 nmol/min/mg cells, and a pH optimum of 5.0; (b) the V for NCAO transport in gap mutants, which lack the general amino acid transport system, was approximately 1% of that observed with wild-type cells; (c) the V for NCAO in cells deprived of glucose was less than 5% of that observed when glucose was present. NCAO was transiently concentrated more than 1000-fold by yeast cells when glucose served as an energy source. The internal pool of NCAO was metabolized by the yeast cells and the products were excreted. When 100 μ m [ 14C]NCAO was incubated with a yeast cell suspension for 8 h, more than 95% of the compound was converted into two ninhydrin-negative excretory products. The effect of NCAO on the growth of yeast cells was determined. Wild-type strains did not grow when 1 m m NCAO was present in the medium. The growth of gap mutants was not inhibited by 1 m m NCAO.