Calcium oxide is an appropriate catalyst for biodiesel production. In this study, an inexpensive and environment-friendly catalyst was used. The waste mussel shell collected in of Persian Gulf coast is a source of calcium carbonate which changes to calcium oxide in calcinations temperatures higher than 950 C. Transesterification reaction was done in the presence of soybean oil, methanol and mussel shell catalyst in a temperature of 60 C. Using response surface methodology (RSM) the effects of different parameters (calcination temperature, catalyst concentration and molar ratio of methanol to oil) were studied. Each parameter was set at three levels labeled as low, medium and high, respectively (calcination temperature of 950, 1000, and 1050 C; catalyst concentration of 6, 9 and 12 wt.%; methanol to oil ratios of 12:1, 18:1 and 24:1). The results of the optimization implies that the calcination temperature of 1050 C, catalyst concentration of 12 wt.%, and methanol to oil ratio of 24:1 provide the maximum values of purity and yield in biodiesel production. The reusability of mussel shell catalyst was studied for five times and the result showed that the catalyst re-calcination in reusability step has negative effects on the yield of biodiesel production.