Computational design, structure refinement and molecular dynamics simulation of novel engineered serratiopeptidase analogs

Novel serratiopeptidase exhibits different affinities to the substrates and inhibitors

“…Considering various analyzed MD simulation parameters (RMSD, R g , and RMSF), all variants exhibited a good stability and acquired a stable and compact conformation similar to the native enzyme during MD simulations (Rouhani et al, 2019). However, in the present study, T306[12‐302ss], T344[8‐339ss], and T380[12‐302ss] variants, which possess 167, 129, and 93 residues lesser than the native enzyme (473 aa), respectively, were selected for docking studies with different truncation lengths.…”

“…In our previous work, different truncations were implemented on the 3D-structure of the native enzyme (PDB ID: 5d7w) using Swiss-PDB (SPDB) viewer v.4.1 (Guex & Peitsch, 1997) and visual molecular dynamics (VMD) v.1.9 (Humphrey et al, 1996). Moreover, A8C-V339C and L12C-R302C double mutations were considered as potential sites for disulfide bond formation to enhance the stability of the enzymes structure (8-339ss and 12-302ss) (Rouhani et al, 2019). The structural stability of the variants was then investigated by MD simulation using GROMACS v.4.6.5 package (Abraham et al, 2015) on a Linux Mint 17 operating system for 100 ns.…”

“…The trajectories were finally analyzed in terms of root mean square deviation (RMSD), radius of gyration (R g ), root mean square fluctuation (RMSF), and structural superimposition. Based on the data, especially fluctuations in the active site residues, T306[12-302ss], T344 [8-339ss], and T380[12-302ss] variants were selected for docking study (Rouhani et al, 2019).…”

“…Serratiopeptidase specifically acts on peptide bonds of glycine-hydrophobic amino acids. Molecular docking studies were carried out using 3D-models generated in our previous study (Rouhani et al, 2019) to analyze the enzyme-substrate interaction mode. In this regard, CABS-dock Web server (Ciemny et al, 2017) was used for substrate docking including Bradykinin (peptide sequence: RPPGFSPFR) and substance-P (peptide sequence: RPKPQQFFGLM).…”

“…In our previous study, six mutated and/or truncated variants of the native enzyme (Serra 473) have been designed and the structural stability was assessed using molecular dynamics (MD) simulation (Rouhani et al, 2019). Based on the results obtained from this study, four engineered forms were expressed as recombinant proteins in Escherichia coli cells, then purified and were subjected to functional assessment.…”