AppA C-terminal Plays an Important Role in its Thermostability in Escherichia coli

Abstract

Due to our previous research, mainly the thermostable mutants Q307D, Y311K, and I427L, we conjectured that Escherichia coli AppA phytase's C-terminal plays an important role in its thermostability, and AppA begins to collapse from the C-terminal when at a higher temperature. So here we constructed C-lose mutant to prove it. The residual activities of the wild-type AppA phytase and C-lose were 31.42 and 70.49 %, respectively, after being heated at 80 °C for 10 min. The C-terminal deletion mutant C-lose showed 39.07 % thermostability enhancement than the wild-type both without the pH and temperature optimum changed. It proved the C-lose plays a key role in E. coli AppA phytase's thermostability.

Insights into the unfolding pathway and identification of thermally sensitive regions of phytase from Aspergillus niger by molecular dynamics simulations

“…The helix was found to be very well conserved in the C-terminus until the simulated temperature rose to 500 K. In general, for most of the proteins, the N-terminus shows more flexibility than the C-terminus. Hence, it can be inferred from our results that either stabilizing or deleting the C-terminus could result in a thermostable variant [15]. The results obtained in this study are qualitative rather than quantitative.…”

“…3c) which contribute greatly to the formation and stability of secondary structures. As the temperature is increased from 310 K to 400 K, the total number of main chain to main chain hydrogen bonds gradually decreases, implying that hydrogen bonding is an important factor in the thermostability of PhyA [12][13][14][15][16][17][18][19]. Also, the solvent-accessible surface area increases significantly at 500 K due to the unfolding of PhyA (Fig.…”

“…A previous study showed that C-terminus deletion can greatly increase the thermal stability of a phytase (AppA in E. coli) [15]. The C-terminus has a destabilizing effect on the α/β fold in AppA, so deleting the C-terminus increased the thermostability of AppA by 39.07 %.…”

Current Progresses in Phytase Research: Three‐Dimensional Structure and Protein Engineering

“…As mentioned in the previous section, the C-terminal residues which form a flexible region were deleted to increase residual activity at 80°C [48]. In another report, the authors identified uncharged residues in the highly flexible areas on the protein surface with molecular dynamic simulation and 3D structures.…”

“…ble 3 10 helix, which is considered prone to collapse at high temperatures. Indeed, deleting the C-terminal region resulted in 39 % higher residual activity at 80°C [48].…”