Study on the Relationship Between Cyclodextrin Glycosyltransferase Thermostability and Salt Bridge Formation by Molecular Dynamics Simulation
Abstract:Cyclodextrin glycosyltransferase (EC 18.104.22.168, CGTase) is an important industrial enzyme in the production of cyclodextrins. Thermal stability is of great importance for this enzyme. Rational design of thermostable variants of mesophilic proteins is well motivated. In this work, molecular dynamics simulations have been performed to study thermal stabilization of CGTase protein via electrostatic interactions of salt bridges. To predict behaviors of the salt bridges engineered into a mesophilic protein to increase stability, in silico mutant of CGTase from the mesophilic Bacillus macerans is generated. Dynamic motions of salt bridges in thermal unstable regions are monitored during the simulations. Among the five salt bridges, Lys88-Glu91, Asp296-Arg335 and Arg336-Asp370 are found to be more important for stability than the others. Especially, the region C is stabilized by a well-organized strong multiple salt bridge interactions. The results reveal that salt bridges involved in thermal unstable regions are relatively strong and prone to be tightened at elevated temperature, which can hold the stable conformation of the spatial neighborhood. Meanwhile, we use the heat capacity and total energy as the measure of stability difference between the original and its mutant variant, and then, quantify the contribution of salt bridges in thermal unstable regions for the mutant protein. Therefore, the viable computational strategy has been demonstrated to improve thermal stability of the mesophilic CGTase by introducing stable salt bridge interactions into its thermal unstable regions and it can be universally applied to other enzymes.
Keywords: (NVT); (PME); (RMSD); (RMSF); (SDM); Arg336-Asp370; Asp296-Arg335; CGTase; CHARMM27; Heat capacity; Lys88-Glu91; MD simulations; NAMD; NMR; Thermostability; bridge; capacity; cyclodextrins; enzyme; heat capacity; low-frequency; meso-philic; molecular; molecular dynamics simulation; mutation; salt bridge; thermal stability; thermophilic
Document Type: Research Article
Publication date: November 1, 2010
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