# Peptide Mediated Antimicrobial Dental Adhesive System
## 3 Results and Discussion
### Computational Structure Flexibility
Peptides are not static when floating in solution, instead they are often modeled as dynamic mechanical systems of springs and masses. constantly in motion [45; 86]. The computational folding model and the circular dichroism measurements agreed with results for increasingly ordered water conditions with TFE in the buffering solution, but not for the less-ordered water conditions with phosphate-buffer only. The computational folding model used an implicit solvent method, which provided a faster structure generation process. Another advantage to the implicit solvent model was the averaging of the solvent conditions of ordered peptide states, which are closely related to the environmental conditions of the fragments when the structure was determined. Figure 4: Analysis of secondary structure features predicted through PyRosetta method and quantified through DSSP(Define Secondary Structure of Proteins). (**a**) GH12 flexibility analysis of percentages of secondary structures (rows) occurring in the PyRosetta-generated decoys by residue calculated by DSSP. (**b**) GH12-M2 flexibility analysis of secondary structure of decoys by residue using PyRosetta and DSSP. (**c**) GH12 lowest energy score structure decoy generated through PyRosetta method. (**d**) GH12-M2 lowest energy structure. Green ribbon indicates the GH12 residues, and the purple ribbon indicates the modification for GH12-M2. This environment was not in disordered conditions such as in a dilute aqueous solution, rather fragments were obtained from crystallized structures in ordered environments. The computational folding model estimated a higher occurrence of helical structures, while the deconvolution of the CD spectra by CD Pro predicted mainly beta-sheet structures for peptides in buffer only. However, the deconvolution method observed large increases in the helical structures with increasing amounts of 2,2,2-trifluoroethanol (TFE) in the solution. TFE is a known kosmotropic agent, increasing the order of the water near the surface of biomolecules [87]. TFE has long been used in the crystallization of proteins. The increased TFE concentrations were expected to put the peptides in an ordered, structured state [88; 89]. The computational folding model held a bias toward native, ordered structures because it used fragments from crystallized structures. Whereas fragment-insertion structures generated were representative of kosmotropic agent-related peptide structures, obtained with the increasing levels of TFE. The method held a bias against generating structures representative of peptide structures with chaotropic agents such as would likely be observed with urea or guanidinium chloride. The water near bacterial surfaces is likely also highly ordered, due to the biomolecules of the bacterial membrane [76; 90]. Since our structure generation method searched the portions of the structural space with ordered fragments, it may provide structures representative of bacterial surface interactions for structure-function relationships. We next explored the functionality of the peptide engineered adhesive formulations on the discs. Incorporation of additional functionality to antimicrobial peptides benefits from detailed analyses of the structure-function relationships on the resulting peptides as discussed in our group's recent papers on antimicrobial peptides based coatings or films [43; 44; 54; 91]. In this study design, we investigated if residues of the spacer domain added for the conjugation to the polymer affect the peptide folding, which in turn may have an adverse effect on the antimicrobial property. We observed similar circular dichroism spectra for GH12 and GH12-M2 in Figure 2. We also obtained similar analyses for both peptides using FTIR and Raman spectra as shown in Figure 3. Almost no difference in structural flexibility in the peptide models were observed (Figure 4).