Strategies for bacterial expression of protein–peptide complexes: Application to solubilization of papillomavirus E6

摘要: Abstract E6 is a small oncoprotein involved in tumorigenesis induced by papillomaviruses (PVs). often recognizes its cellular targets binding to short motifs presenting the consensus LXXLL. proteins have long resisted structural analysis. We found that bovine papillomavirus type 1 (BPV1) binds N-terminal LXXLL motif of protein paxillin with significantly higher affinity as compared other E6/peptide interactions. Although recombinant BPV1 was poorly soluble free state, provision peptide during biosynthesis greatly enhanced protein’s solubility. Expression E6/LXXLL complexes carried out bacteria form triple fusion constructs comprising, from N- C-terminus, carrier maltose (MBP), and protein. A TEV protease cleavage site placed either between MBP or E6. These allowed us produce highly concentrated samples E6, covalently fused C-terminus (intra-molecular complex) non-covalently bound it (inter-molecular complex). Heteronuclear NMR measurements were performed showed folded similar conformations both covalent non-covalent complexes. data open way novel functional studies complex preferential target motif.