Evaluating the strengths of salt bridges in the CutA1 protein using molecular dynamic simulations: a comparison of different force fields

Ion–ion interactions (salt bridges) between favorable pairs of charged residues are important for the conformational stability of proteins. Molecular dynamic (MD) simulations are useful for elucidating the interactions among charged residues fluctuating in solution. However, the quality of MD results depends strongly on the force fields used. In this study, we compared the strengths of salt bridges among force fields by performing MD simulations using the CutA1 protein (trimer) from the hyperthermophile Pyrococcus horikoshii (PhCutA1), which has an unusually large proportion of charged residues. The force fields Chemistry at HARvard Macromolecular Mechanics (Charmm)27, Assisted Model Building and Energy Refinement (Amber)99sb, Amber14sb, GROningen Molecular Simulation (Gromos)43a1, and Gromos53a6 were used in combination with two different water models, tip3p (for Charmm27, Amber99sb, and Amber14sb) and simple point charge/extended (for Amber99sb, Gromos43a1, and Gromos53a6), yielding a total of six combinations. The RMSDs of all Cα atoms of PhCutA1 were similar among force fields, except for Charmm27, during 400‐ns MD simulations at 300 K; however, the radius of gyration (R g) was greater for Amber99sb and shorter for Gromos43a1. The average strengths of salt bridges for each positively charged residue did not differ greatly among force fields, but the strengths at specific sites within the structure depended sensitively on the force field used. In the case of the Gromos group, positively charged residues could engage in favorable interactions with many more charged residues than in the other force fields, especially in loop regions; consequently, the apparent strength at each site was lower.

. 60 inter-subunit interactions between favorable ion pairs in PhCutA1 force fields Charmm27_tip3p 77.0 ± 1.2 39.4 ± 0.7 33.3 ± 0.6 Amber99sb_spc/e 77.5 ± 1.5 39.5 ± 0.8 31.7 ± 1.4 Amber99sb_tip3p 76.6 ± 1.4 38.7 ± 0.9 30.9 ± 1.5 Amber14sp_tip3p 78.1 ± 0.9 39.9 ± 0.5 33.4 ± 0.7 Gromos43a1_spc/e 76.6 ± 2.0 38.0 ± 1.7 33.6 ± 0.5 Gromos53a6_spc/e 75.6 ± 1.8 37.3 ± 1.4 33.6 ± 0.4 *structure = β-sheet + α-helix + β-bridge + turn Table S2. Number of residues of PhCutA1 in each type of secondary structure in MD simulations (50-400 ns) structure* β-sheet α-helix Values represent the average number of residues in each type of secondary structure among three subunits. These values represent the average of three subunits. All distances are in nm. These data are shown when ion-pairs less than 0.7 nm were detected.at least once among six force fields. Yellow and orange represent, respectively, the lowest and highest values of the distance among six force fields. All distances are in nm. A, B, and C represent the A-, B-, C-subunits of PhCutA1, respectively These data are shown when ion-pairs less than 0.7 nm were detected.at least once among six force fields. Yellow and orange represent, respectively, the lowest and highest values of the distance among six force fields.  Table S6. Comparison of percent occupancy of intra-subunit salt bridges in each subunit of PhCutA1 during 400-ns MD simulation at 300 K using indicated force fields Data show average values of percent occupancies of 17 positively charged residues indicated in Table 2A.
STDEV represent the the standard deviation of average values for 3 subunits. The side chains of charged residues in PhCutA1 shown in Figure S4 to Figure S14 were examined by MolProbity (http://molprobity.biochem.duke.edu).      Targeted ionic pairs are listed in Tables S1A and S1B for intra-and inter-subunit interaction, respectively. Three bars represent the data for each targeted pair in A, B, and C-subunits. The     Green, cyan, and magenta represent A, B, and C-subunits of PhCutA1, respectively.
Green, cyan, and magenta represent A, B, and C-subunits of PhCutA1, respectively.
(A) The crystal structure of PhCutA1 (A, B, and C-subunits of 4nyo).
(B) The snapshot of PhCutA1 at 200 ns of an MD simulation in the case of Gromos43a1_spc/e. Figure S15. Trajectories of distance between Arg33 in PhCutA1 and Clion during 400-ns MD simulations at 300 K using indicated force fields (a) Charmm27_tip3p, the distance between C ζ of Arg33 in A-subunit and Clion of the number 12197. The percent occupancy of distance (less than 0.6 nm) between them was 100.0 %.
(b) Amber99sb_tip3p, the distance between C ζ of Arg33 in A-subunit and Clion of the number 12197. The percent occupancy of distance (less than 0.6 nm) between them was 86.0 %.
(c) Gromos43a1_spc/e, the distance between C ζ of Arg33 in C-subunit and Clion of the number 12220. The percent occupancy of distance (less than 0.6 nm) between them was 1.7 %.