Structural Insights into Antibacterial Payload Release from Gold Nanoparticles Bound to E. coli Peptide Deformylase

Abstract

The lack of new antibiotics and the rapidly rising number of pathogens resistant to antibiotics pose a serious problem to mankind. In bacteria, the cell membrane provides the first line of defence to antibiotics by preventing them from reaching their molecular target. To overcome this entrance barrier, it has been suggested[1] that small Gold‐Nanoparticles (AuNP) could possibly function as drug delivery systems for antibiotic ligands. Using actinonin‐based ligands, we provide here proof‐of‐principle of AuNP functionalisation, the capability to bind and inhibit the target protein of the ligand, and the possibility to selectively release the antimicrobial payload. To this end, we successfully synthesised AuNP coated with thio‐functionalised actinonin and a derivative. Interactions between 15N‐enriched His‐peptide deformylase 1–147 from E. coli (His‐ecPDF 1–147) and compound‐coated AuNP were investigated via 2D 1H‐15N‐HSQC NMR spectra proving the direct binding to His‐ecPDF 1–147. More importantly by adding dithiothreitol (DTT), we show that the derivative is successfully released from AuNPs while still bound to His‐ecPDF 1–147. Our findings indicate that AuNP‐conjugated ligands can address and bind intracellular target proteins. The system introduced here presents a new delivery platform for antibiotics and allows for the easy optimisation of ligand coated AuNPs.

Toward More Selective Antibiotic Inhibitors: A Structural View of the Complexed Binding Pocket of E. coli Peptide Deformylase

“…Notably, the observed shifts indicate a fast-exchange and were small in comparison with other compounds like actinonin and some of its derivatives. 39 Taken together, only for backbone amide resonances of 12Arg, 43Gly, 63Ser, 91Leu, 92Ser, 100Val, 102Arg N η -H η (side chain), 105Lys, 129Cys, 130Ile, and 147Ser shifts and secondary signals could be detected. Notably, 86Ile and 90Cys did not show any secondary signal but a shifting vector with changing directions.…”

“…Resonances from 41Glu, 42Glu, 46Leu, 50Gln, 66Arg, 67Asp, 78Leu, 87Glu, 88Glu, 115Gly, 133Glu, 134Met, 139Glyamong othersshowed small shifts (below SD, see the CSP plot) and no secondary signals. Notably, the observed shifts indicate a fast-exchange and were small in comparison with other compounds like actinonin and some of its derivatives …”

“…The isolation procedure of ecPDF full length (ecPDF fl) and ecPDF 1-147 was performed as previously published. 39 It was adopted from Meinnel et al 51 and Byerly et al 45 Briefly, the corresponding protein sequence was cloned into a modified pET19 vector with an additional N-terminal TEV-protease signal sequence between the His 6 -tag and the protein sequence by restriction and ligation. For His-ecPDF 1-147 used in NMR experiments, competent BL21 (DE3) RIL cells were transformed and used to inoculate 250 mL of the 12 C− 14 N-M9 minimal medium.…”