دانلود رایگان مقاله بینش به تاثیر دو یون کلسیم ساختاری بر خواص قارچ ارینجی

Abstract

Two structural Ca2+ (proximal and distal) is known to be important for ligninolytic peroxidases. However, few studies toward impact of residues involved in two Ca2+ on properties of ligninolytic peroxidases have been done, especially the proximal one. In this study, mutants of nine residues involved in liganding two Ca2+ of Pleurotus eryngii versatile peroxidase (VP) were investigated. Most mutants almost completely lost activities, except the mutants of proximal Ca2+ - S170A and V192T. In comparison with WT (wild type), optimal pH values of S170A, S170D, and V192T shifted from pH 3.0 to pH 3.5. The order of thermal and pH stabilities of WT, V192T, S170A, and S170D is similar to that of their specific activities: WT > V192T > S170A > S170D. The CD (circular dichroism) results of WT and several mutants indicated that mutations had some effects on secondary structures. For the first time, it was observed that the thermostability of ligninolytic peroxidases is related with proximal Ca2+ too, and the mutant containing distal Ca2+ only was obtained. Our results clearly demonstrated that enzymatic activities, pH and thermal stabilities, Ca2+content, and secondary structures of VP have close relationship with the residues involved in two structural Ca2+.

5. Conclusion

Ten mutants of the residues liganding two structural calcium ions of VP were made and fully characterized. Most mutations resulted in great loss of enzyme activity. The different impact of residues involved in two calcium ions on enzyme activity, UVevis spectroscopy, and secondary structure was observed, and the effects of the mutants related to proximal Ca2þ only on enzyme activity, UVevis spectroscopy, pH and thermal stability, and circular dichroism were presented for the first time. It was observed that the thermostability of ligninolytic peroxidases such as VP is related with proximal Ca2þ for the first time as well. Our results confirmed the significance of residues coordinating two structural calcium ions of ligninolytic peroxidases and close relationship between enzyme activity and Ca2þ concentration in protein, and would provide a guide for engineering them towards higher thermal and pH stability.