Structural highlights
Publication Abstract from PubMedPglK is an ABC transporter that flips a lipid-linked oligosaccharide (LLO) that serves as a donor in protein N-glycosylation. Previous structures revealed two inward-facing conformations, both with very large separations of the nucleotide binding domains (NBDs), and a closed, ADP-bound state that featured an occluded cavity. To investigate additional states, we developed conformation-sensitive, single-domain camelid nanobodies (Nb) and studied their effect on PglK activity. Biochemical, structural, and mass spectrometric analyses revealed that one inhibitory Nb binds as a single copy to homodimeric PglK. The co-crystal structure of this Nb and ADP-bound PglK revealed a new, narrowly inward-open conformation. Rather than inducing asymmetry in the PglK homodimer, the binding of one Nb results in steric constraints that prevent a second Nb to access the symmetry-related site in PglK. The Nb performed its inhibitory role by a "sticky-doorstop" mechanism, where inhibition of ATP hydrolysis and LLO flipping activity occurs due to impaired closing of the NBD interface, which prevents PglK from converting to an outward-open conformation. This inhibitory mode suggests tight conformational coupling between the ATPase sites, which may apply to other ABC transporters. Structural basis of inhibition of lipid-linked oligosaccharide flippase PglK by a conformational nanobody.,Perez C, Kohler M, Janser D, Pardon E, Steyaert J, Zenobi R, Locher KP Sci Rep. 2017 Apr 19;7:46641. doi: 10.1038/srep46641. PMID:28422165[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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