Disorder is a critical component of lipoprotein sorting in Gram-negative bacteria

We thank Asma Boujtat for technical help. We are indebted to the members of the Collet laboratory and to Nassos Typas (EMBL, Heidelberg) for helpful suggestions and discussions and to Tom Silhavy (Princeton) for providing bacterial strains. J.S. was a research fellow of the FRIA and J.F.C. is an Investigator of the FRFS-WELBIO. This work was funded by the WELBIO, by grants from the F.R.S.-FNRS, from the Fédération Wallonie-Bruxelles (ARC 17/22-087), from the European Commission via the International Training Network Train2Target (721484), and from the EOS Excellence in Research Program of the FWO and FRS-FNRS (G0G0818N).; Gram-negative bacteria express structurally diverse lipoproteins in their envelope. Here we found that approximately half of lipoproteins destined to the Escherichia coli outer membrane display an intrinsically disordered linker at their N-terminus. Intrinsically disordered regions are common in proteins, but establishing their importance in vivo has remained challenging. Here, as we sought to unravel how lipoproteins mature, we discovered that unstructured linkers are required for optimal trafficking by the Lol lipoprotein sorting system: linker deletion re-routes three unrelated lipoproteins to the inner membrane. Focusing on the stress sensor RcsF, we found that replacing the linker with an artificial peptide restored normal outer membrane targeting only when the peptide was of similar length and disordered. Overall, this study reveals the role played by intrinsic disorder in lipoprotein sorting, providing mechanistic insight into the biogenesis of these proteins and suggesting that evolution can select for intrinsic disorder that supports protein function.