An experiment was conducted to examine how the response of dairy cows to a change from twice to three times-daily milking is affected by deficiencies in the dietary supplies of three amino acids, His, Met, and Lys. Six cows were used in a 6 x 6 Latin square with 14-d periods. The three dietary treatments were: grass silage and a cereal-based supplement containing feather meal as the sole protein supplement; the same silage-cereal diet supplying similar amounts of metabolizable and rumen-undegradable protein but with additional amounts of His, Met, and Lys in the form of fish meal; and the fish meal diet with additional metabolizable energy in the form of an additional 2 kg/d of sugar beet pulp. Within each of these dietary treatments, the cows were milked twice and three times daily, making a total of six treatments. When cows were given the feather meal diet, even though dietary metabolizable energy was in considerable excess, a deficiency of specific amino acids prevented any increase in milk yield in response to increasing the frequency of milking from twice to three times daily. In contrast, when cows consumed a similar level of excess metabolizable energy and a similar level of rumen-undegradable protein for which the protein was of better amino acid balance (fish meal), the increased frequency of milking led to increased yield of milk and milk protein.