Multiplicity and regulation of amino acid transport in Penicillium chrysogenum

Abstract

Penicillium chrysogenum possesses a highly specific sulfur-regulated permease for l-methionine (Benko, Wood, and Segel, Arch. Biochem. Biophys. 122, 783–804 (1967)). In addition, the mycelium has several other relatively specific amino acid permeases. For example, l-phenylalanine- 14C transport by sulfur- and nitrogen-sufficient mycelium is not significantly inhibited by a 10-fold excess of unlabeled l-methionine, l-leucine, l-serine, or l-α-aminobutyric acid. Similarly, l-leucine- 14C transport is not significantly inhibited by a 10-fold excess of any of the other four amino acids. Nitrogen-starvation results in the development of a nonspecific amino acid permease ( V max for l-methionine, l-leucine, and l-phenylalanine ca. 10 μmoles/g-min). The development is prevented by actidione. Structural requirements for interaction with the nonspecific amino acid permease are: (a) an unsubstituted l-α-amino group, (b) a single α-hydrogen atom, (c) no secondary negatively charged group, and (d) an α-carbonyl group. The K m values of the nonspecific permease for l-phenylalanine and l-methionine are ca. 10 −5 m. The K i for l-leucine (with l-methionine as substrate) is 2.4 × 10 −5 m; the K i for l-methionine (with l-leucine as substrate) is 1.4 × 10 −5 m. Other characteristics of the permease are pH-dependence (optimum at 6), temperature-dependence (Q 10 of 2–5 between 15 and 32 °), and energy-dependence (DNP and azide sensitive). The general amino acid permease is independent of monovalent and divalent metal ions and of concurrent protein synthesis for activity. Ammonia (NH 4 +) was the only non-amino acid inhibitor of the permease. The permease activity can be markedly and rapidly reduced by preloading the nitrogen-deficient mycelium with NH 4 + or any one of several amino acids. The results suggest that the nonspecific amino acid permease is subject to feedback regulation by intracellular NH 4 + and amino acid substrates.