Active S68 and inactive SIRS pinholin interact differently with the lipid bilayer: A P and H solid state NMR study.

Pinholins are a family of lytic membrane proteins responsible for the lysis of the cytosolic membrane in host cells of double stranded DNA bacteriophages. Protein-lipid interactions have been shown to influence membrane protein topology as well as its function. This work investigated the interactions of pinholin with the phospholipid bilayer while in active and inactive confirmations to elucidate the different interactions the two forms have with the bilayer. Pinholin incorporated into deuterated DMPC-d lipid bilayers, along with P and H solid state NMR (SS-NMR) spectroscopy were used to probe the protein-lipid interactions with the phosphorus head group at the surface of the bilayer while interactions with the H nuclei were used to study the hydrophobic core. A comparison of the P chemical shift anisotropy (CSA) values of the active S68 pinholin and inactive SIRS pinholin indicated stronger head group interactions for the pinholin in its active form when compared to that of the inactive form supporting the model of a partially externalized peripheral transmembrane domain (TMD) of the active S68 instead of complete externalized TMD1 as suggested by Ahammad et al. JPC B 2019. The H quadrupolar splitting analysis showed a decrease in spectral width for both forms of the pinholin when compared to the empty bilayers at all temperatures. In this case the decrease in the spectral width of the inactive SIRS form of the pinholin showed stronger interactions with the acyl chains of the bilayer. The presence of the inactive form's additional TMD within the membrane was supported by the loss of peak resolution observed in the H NMR spectra.