In order to understanding proteomic basis of drought tolerance in sunflower (Helianthus annuus L.), two contrasting inbred lines were subjected to drought stress during the flowering stage for two years. Proteins were extracted from leaves of well-watered and drought-treated plants by using the TCA–acetone precipitation method and analysed by two-dimensional polyacrylamide gel electrophoresis followed by nanoscale liquid chromatography coupled to tandem mass spectrometry for identification of affected proteins. When comparing proteomic patterns, 18 proteins were changed by drought stress in sensitive lines and 24 proteins in tolerant lines. Concurrent down-expressions of oxygen-evolving enhancer and ferredoxin-NADP reductase were considered as primary drought sensors that mediate downstream pathways to cope with drought conditions. Differential and line-specific proteomic changes were attributed as the source for contrasting response to drought stress. According to the results, scavenging of reactive oxygen species, conservation of energy and water, and cell-structure integrity constituted the major aspects of drought tolerance in sunflower.