GENETIC AND AMINO-ACID-ANALYSIS OF 2 MAIZE THREONINE OVERPRODUCING, LYSINE-INSENSITIVE ASPARTATE KINASE MUTANTS

The aspartate-derived amino-acid pathway leads to the production of th e essential amino-acids lysine, methionine, threonine and isoleucine. Aspartate kinase (AK) is the first enzyme in this pathway and exists i n isoforms that are feedback inhibited by lysine and threonine. Two ma ize (Zea mays L.) threonine-overproducing, lysine-insensitive AK mutan ts (Ask1-LT19 and Ask2-LT20) were previously isolated. The present stu dy was conducted to determine the map location of Ask2 and to examine the amino-acid profiles of the Ask mutants. The threonine-overproducin g trait conferred by Ask2-LT20 was mapped to the long arm of chromosom e 2. Both mutants exhibited increased free threonine concentrations (n mol/mg dry weight) over wild-type. The percent free threonine increase d from approximately 2% in wild-type kernels to 37-54% of the total fr ee amino-acid pool in homozygous mutant kernels. Free methionine conce ntrations also increased significantly in homozygous mutants. Free lys ine concentrations were increased but to a much lesser extent than thr eonine or methionine. In contrast to previous studies: free aspartate concentrations were observed to decrease, indicating a possible limiti ng factor in threonine synthesis. Total (free plus protein-bound) amin o-acid analyses demonstrated a consistent, significant increase in thr eonine, methionine and lysine concentrations in the homozygous mutants . Significant increases in protein-bound (total minus free) threonine, methionine and lysine were observed in the Ask mutants, indicating ad equate protein sinks to incorporate the increased free amino-acid conc entrations. Total amino-acid contents (nmol/kernel) were approximately the same for mutant and wild-type kernels. In five inbred lines both Ask mutations conferred the threonine-overproducing phenotype, indicat ing high expressivity in different genetic backgrounds. These analyses are discussed in the context of the regulation of the aspartate-deriv ed amino-acid pathway.