Thermostable chitinases are useful for industrial and biotechnological applications. This paper reports the stabilization of chitinase from Serratia marcescens B4A through rational mutagenesis. Changing of Ser 390 to Ile in S. marcescens. The stabilization was enhanced through entropic stabilization by reduction of the loop length and also by increasing of the beta chain length. With this replacement, polar uncharged residue changed to non-polar one and increased the hydrophobic interactions. Furthermore Isoleucine has branched β-carbon that restricts the backbone conformation more than non-branched residues. Finally all of these factors lead to entropic stabilization and thermal stabilization. The results exhibited that the optimal temperature and pH for enzyme activity of native chitinase were not changed by mutagenesis which showed that mutation didn’t affect the original characteristics of the enzyme, the K_m values of native and mutant chitinase were different very little, showing that the affinity of enzyme towards the substrate and also the natural flexibility of chitinase did not change by mutation. Besides the V_max value of the mutant chitinase was decreased, while its pH stability was increased briefly, but its thermal stability was increased remarkably. Mutation made chitinase to tolerate high temperatures up to 90^°C. In addition its activity was increased at 50^°C, 60^°C for 120 min and up to 2 hours of incubation period and the mutant chitinase demonstrated a high level of activity at 60^°C. These results show that entropic stabilization works well for chitinase and this approach may be generally applicable for stabilization of other proteins.