Genetic alterations of ribonuclease-sensitive glycoprotein subunits of amino acid transport systems in Neurospora crassa conidia

Abstract

Neurospora crassa conidia have multiple and constitutive amino acid transport systems. Extraction by KCl releases amino acid-binding glycoproteins which have been purified by arginine affinity chromatography. Disappearance of certain fractions is coordinate with genetic lesions which reduce amino acid transport. Two such affinity fractions contain radioactivity when cells are grown on l-[ 14C]phenylalanine or on [ 14C]uridine, but not when cells are grown on [ 14C ]glucosamine. One purified arginine-binding fraction (B) contains 113 amino acid residues per minimum molecular weight. This glycoprotein also contains eight types of neutral sugar residues. No amino sugars were detected. Electrophoresis of crude extracts reveals five major Coomassie blue-staining species. The number of species is reduced, and the electrophoretic pattern is altered in extracts from transport-deficient strains. Tryptic “fingerprints” of these extracts indicate that mutations that reduce transport result in amino acid substitutions in the extractable glycoproteins. Nondialyzable material which absorbs light in the 260-nm region becomes dialyzable after digestion with RNase. Digestion of conidia with RNase reduces the amount of l-phenylalanine accumulated by the cells after 10 min of incubation with the amino acid.