Comparative Study Of The Hydrolase Activities Of Myoglobin And Its Mutants

Heme proteins perform a variety of biological functions in living systems, which are important metalloproteins. Although heme proteins have been extensively studied with fruitful achievments, the relationship between the structure and hydrolase function of heme proteins is still not fully understood. In this study, we compared the structure of heme distal site of wild type myoglobin(WT Mb) and its two mutants L29 H Mb and L29 E Mb, and found that the hydrolytic activity of L29 E Mb was higher than that of WT Mb and L29 H Mb. Fluoride ions(F-) competition experiment showed that the axial water molecule of L29 E Mb was more likely to be replaced by F- than other proteins, whereas the axial water molecule of L29 H Mb was more stable. p H titration study showed that the p Ka of L29 E Mb was lower than that of the WT Mb, whereas L29 H Mb’s p Ka was higher than that of the WT Mb. In addition, the p H-jump study showed that the distal heme hydrogen bond network played a key role to the stability of the protein. These findings suggested that the hydrogen-bonding network in the heme distal site as formed by the microenvironment regulateed the protein structure and hydrolysis activity. In conclusion, this study provides important information for understanding the relationship between the structure and hydrolysis function of heme proteins, moreover, it provided a new direction for design of functional proteins.