The role of electrostatic interactions on klentaq1 insight for domain separation

Abstract

We investigated the relationship between the thermostability of Klentaq1 and factors stabilizing interdomain interactions. When thermal adaptation of Klentaq1 was analyzed at the atomic level, the protein was stable at 300 and 350 K. It gradually unfolded at 373 K and almost spontaneously unfolded at 400 K. Domain separation was induced by disrupting electrostatic interactions in two salt bridges formed by Lys354-Glu445 and Asp371-Arg435 on the interface domain. The role of these interactions in protein stability was evaluated by comparing free energy solvation (ΔΔG(solv)) between wild type and mutants. Substitution of Asp371 by Glu or Asn, and also Glu445 by Asn resulted in a positive value of ΔΔG(solv), suggesting that mutations destabilized the protein structure. Nevertheless, substitution of Glu445 by Asp gave a negative value to ΔΔG(solv) reflecting increasing protein stability. Our results demonstrate that interactions at the interface domains of Klentaq1 are essential factors correlated with the Klentaq1 thermostability.

Keywords: Klentaq1; domain separation; electrostatic interaction; in silico mutation.

Figure 1

Root-mean-square deviation (RMSD) values of the Cα of Klentaq 1 at various temperatures as a function of simulation time. Notes: Blue line, RMSD at 300 K; Red line, RMSD at 350 K; Green line, RMSD at 373 K; and Purple line, RMSD at 400 K.

Figure 2

Thermostability map of Klentaq1 based on B-factor value for 300 K simulation. Notes: The thermostability map was illustrated in RGB (Red Green Blue) color scale, where the rigid (stable) to flexible region is showed as a gradation in color from red to blue. The backbone atoms are displayed as a new ribbon to show the secondary structure (Drawn with VMD).

Figure 3

(A) Percentage of secondary structure for Klentaq1 during 373 K simulation, α-helix (blue line) and β-sheet (red line). (B) Solvent accessible surface area (SASA) values for the total area (blue line), polar region (red line), non-polar region (green line) and backbone (purple line) of Klentaq1 during 373 K simulation.

Figure 4

Conformational changes in Klentaq1 observed in the unfolding profile at 373 K simulation. Note: The circles indicate early events in the unfolding process (Drawn with VMD).

Figure 5

The distance of electrostatic interaction (salt bridges) ie, Asp371-Arg435 (blue line) and Lys354-Glu445 (red line) during 373 K simulation.