# Incidence of Phage Capsid Organization on the Resistance to High Energy Proton Beams
## 3 Results and Discussion
Specifically, 12 III1 foreign peptide contains amino acids with aromatic (4), negatively (3) and positively (3) R-groups that are highly responsive to oxidation products from water radiolysis, such as with OH\({}^{-}\) addition to the aromatic ring or carbonylation reaction. Moreover, oxidation by reactive oxygen species (ROS) could also include the production of carbonyl groups on the amino acid side \begin{table}
\begin{tabular}{c c c c c} \hline  & & **Dose (Gy)** & & \\ \hline
50 & 125 & 250 & 375 & 500 \\ \hline  & & **Dose Rate (Gy/s)** & & \\ \hline
0.0615 & 0.155 & 0.309 & 0.463 & 0.615 \\ \hline \end{tabular}
\end{table} 
Table 1: Used dose and dose rates for phage solution proton irradiation. Figure 2: Proton penetration in the water solution (**a**) and stopping power of the protons along the solution depth (**b**). chain of proline residues. On the other hand, in P9b, the presence of aliphatic amino acids, namely leucine (2) and alanine (2), in foreign peptide should maintain the capsid's integrity and, therefore, the phage clone is less susceptible to oxidative stress. All experiments were carried out using M13 wild-type vector without the peptide insert (pC89) as control. Results on the viability of pC89, P9b and 12 III1 phages after exposure to proton beam radiation are reported in Table 2.  
Data show that at the maximum absorbed proton dose (500 Gy) the most resistant phage was P9b (0.38% of survival), while the least resistant was 12 III1 (0.00047%). Finally, an intermediate level of resistance was exhibited by the pC89 (0.033%). This trend of susceptibility was observed at each dose Gy, and P9b was always the most resistant phage, while the least was 12 III1 (Figure 3a). Since the tested phages have the same DNA, with the exception of the small insert displaying the foreign peptide on the major coat proteins, it is very likely that the difference in susceptibility is due to the presence/absence and amino acid composition of the foreign peptide. The latter does not affect the replication of the phage but determines its susceptibility in the presence of radiation stress. Moreover, since the experimental plan for each phage was carried out in about 800 s continuum irradiation time, the same trend was \begin{table}
\begin{tabular}{c c c c c c c} \hline \hline \multirow{2}{*}{**Phage Clone**} & \multirow{2}{*}{**Exposed**} & \multicolumn{4}{c}{**Dose (Gy)**} \\ \cline{3-6}  & & **50 Gy** & **125 Gy** & **250 Gy** & **375 Gy** & **500 Gy** \\ \hline \multirow{2}{*}{pC89} & (3.0 \(\pm\) 0.14) \(\times\) 10\({}^{10}\) & (2.1 \(\pm\) 0.3) \(\times\) 10\({}^{10}\) & (7.1 \(\pm\) 0.27) \(\times\) 10\({}^{8}\) & (3.0 \(\pm\) 0.41) \(\times\) 10\({}^{8}\) & (1.1 \(\pm\) 0.8) \(\times\) 10\({}^{8}\) & (1.0 \(\pm\) 0.5) \(\times\) 10\({}^{7}\) \\  & (100\%) & (70\%) & (2.37\%) & (1\%) & (0.37\%) & (0.033\%) \\ \hline P9b & (2.6 \(\pm\) 0.21) \(\times\) 10\({}^{10}\) & (2.0 \(\pm\) 0.25)\(\times\)10\({}^{10}\) & (1.1 \(\pm\) 0.17) \(\times\) 10\({}^{10}\) & (5.0 \(\pm\) 0.34) \(\times\) 10\({}^{9}\) & (2.4 \(\pm\) 0.45) \(\times\) 10\({}^{9}\) & (1.0 \(\pm\) 0.49) \(\times\) 10\({}^{8}\) \\ (QRKLAAKLT) & (100\%) & (77\%) & (42.3\%) & (19.2\%) & (9.2\%) & (0.38\%) \\ \hline \multirow{2}{*}{12 III1 (RWPPHFEWHFD)} & (3.0 \(\pm\) 0.38) \(\times\) 10\({}^{10}\) & (5.1 \(\pm\) 0.23) \(\times\) 10\({}^{8}\) & (3.0 \(\pm\) 0.39) \(\times\) 10\({}^{7}\) & (1.9 \(\pm\) 0.5) \(\times\) 10\({}^{7}\) & (4.7 \(\pm\) 0.48) \(\times\) 10\({}^{6}\) & (1.4 \(\pm\) 0.51) \(\times\) 10\({}^{4}\) \\  & (100\%) & (17\%) & (0.1\%) & (0.063\%) & (0.015\%) & (0.00047 \(\times\) 10\%) \\ \hline \hline \end{tabular}
\end{table} 
Table 2: Active virus particles of pC89, P9b and 12 III1, expressed as transducing unit per milliliter of infected _E. coli_ (TU/mL), at different dose (Gy) of 5 MeV proton irradiation. In brackets are survival percentages compared to exposed phages. Each value has been derived as averages from 5 replicates with standard deviations.