Clostridium thermocellum, an anaerobic, thermophilic, and cellulolytic bacterium, is an attractive ethanogenic organism for renewable biomass conversion. However, its low ethanol tolerance has been an obstacle for the process industrialization. A strain tolerant of 4% (w/w) ethanol obtained by serial transfer was capable of producing twice higher concentration of ethanol than wild type. Transcriptomic and proteomic analysis were performed on exponentially growing and stationary phase wild type and tolerant strain. Wild type and tolerant strain exhibited similar dynamics in transiting from growth to stationary phase in many gene classes including cellulosome, glycolysis and protein folding. Whereas many genes involved in membrane transport underwent different responses between them. Combining transcriptomic data with iTRAQ-based quantitative proteomic results, genes and gene classes related to high ethanol producing feature emerged as potential targets for genetic manipulation.