AIMS: To investigate the effect that environmental factors have on Clostridium cellulovorans cellulose binding domain (CBD) binding to a semi-crystalline cellulose ligand, namely Avicel. METHODS AND RESULTS: The behaviour of a 58 kDa mini-CbpA protein containing the CBD from the scaffoldin protein of C. cellulovorans was studied in the presence of various environmental factors, in order to determine whether such factors promote or reduce CBD binding to its ligand, thus potentially affecting its activity on the substrate. The amount of binding was found to be dependent on the Avicel concentration and optimal binding occurred when the ligand concentration was 15 mg ml(-1). Optimal CBD binding occurred at pH 7.0 and at an incubation temperature of 28 degrees C. The effects of dithiothreitol (DTT), 2-mercaptoethanol, acetone, butanol, ethanol and butyric acid were also investigated. CONCLUSIONS: Temperature, pH, DTT, 2-mercaptoethanol and solvents were shown to affect the binding of C. cellulovorans CBD to Avicel. SIGNIFICANCE AND IMPACT OF THE STUDY: Clostridium cellulovorans CBD binding to Avicel is affected by physical conditions and chemicals.