Characterization of Recombinant Chloroperoxidase, and F103A and C29H/C79H/C87H Mutants

Investigating the Role of the Proximal Cysteine Hydrogen Bonding Network and Distal Pocket in Chloroperoxidase

“…as the presence of a sixth ligand, which competes with the exogenous ligand. This explanation correlates with a previous study in our lab, that the CPO mutant, F103A-CPO, also had a higher Kd of 4.5 mM for cyanide [124]. Interestingly, the double, F103A/F186A, mutant displayed a Kd of 3.6 mM which is lesser than that of the two single mutation mutants.…”

“…and 48%, respectively [124]. The different degree of influence between Phe103 and Phe186 might possibly be due to the flexibility of Phe103, as the crystal structure indicates the displacement of Phe103 during the binding of CPD, while Phe186 remained intact [70].…”

“…No sign of chlorination and dismutation activities, increased in epoxidation and peroxidation activities [124] H105A Increased in epoxidation activity, decreased in chlorination and dismutation activity [123] E183H Increased in epoxidation activity, decreased in chlorination and dismutation activities [125] F186A R26A/N33A No sign of chlorination and dismutation activities, increased in epoxidation and peroxidation activities [126] CHAPTER II.…”

“…The result from our lab indicated that the distal pocket of CPO has a dramatic effect on the enzymatic activities as the mutant, F103A-CPO, displayed at least 4-fold enhancement of peroxygenase activity at the expense of the catalase activity. In term of enantioselectivity, F103A-CPO catalyzes epoxidation of long-side chain styrene derivatives with greater enantioselectivity relative to WT-CPO[124]. This mutant revealed the importance of Phe103 on the enantioselective reactions catalyzed by CPO.Phe103 is flexible in the substrate-binding pocket, as the binding of CPD to the active site of CPO resulted in the displacement of Phe103 by 0.5Ǻ away from the active site to accommodate the substrate[70].…”