Levels of ribulose bisphosphate carboxylase (Rubisco) and Rubisco activase were compared in leaves above the ear in two genetically related populations (Z0 and Z20) of maize (Zea mays L.). Z20 was obtained from Z0 after twenty agronomic selection cycles for grain yield improvement (c. 90% above Z0). Plants were cultivated in the highlands of Mexico and leaves were sampled weekly during the grain-filling period. Chlorophyll, soluble protein and Rubisco activity were measured. Chlorophyll and soluble protein content slowly decreased during this period, the former faster than the latter, with no significant differences between populations. During the first 40 days after anthesis, Rubisco activity was significantly greater in the high-yielding population (Z20), although Western blot analysis of Rubisco showed similar values for both populations within this period. However, the same analysis for Rubisco activase indicated a greater amount of this protein in the higher-yielding population (Z20) than the original one (Z0) during the early and middle part of the grain-filling period. The addition of Rubisco activase and an ATP-generating system to Z0 leaf extracts resulted in increased Rubisco activity. It was concluded that during grain-fill in maize, the level of Rubisco activase has a regulatory effect on Rubisco activity expression.