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The opaque2 (o2) mutation in maize (Zea mays L.) increases the content of free amino acids (FAA) in the endosperm. We investigated the basis of this trait by using recombinant inbred lines from a cross of Oh545o2 (high FAA) and Oh51Ao2 (low FAA) to identify quantitative trait loci (QTL) for FAA content and to determine their effect on FAA composition and protein accumulation. Mapping identified six QTLs that accounted for 71% of the phenotypic variation. Two QTLs in bins 4.01 and 7.02 are close to α-zein genes; high FAA individuals with these QTLs had reduced accumulation of α-zein 19 kDa isoforms and increased FAA abundant in α-zeins. A QTL in bin 3.03 is close to a gene encoding triose phosphate isomerase (tpi4) and a higher expression of this enzyme was found in high FAA individuals. Other differentially expressed proteins included vicilin-like globulins and the enzymes glyceraldehyde-3-phosphate …