Available online 7 September 2022, 107896

MlaC comprises two domains viz. NTF2-like and PBP.

Each domain can be divided into two subdomains that are arranged in discontinuous fashion.

MlaC follows a unique “segmented domain movement’’ mechanism.

MlaC has a unique evolutionary ancestry which is different from canonical SBPs.

The outer membrane (OM) of Gram-negative bacteria acts as a formidable barrier against a plethora of detrimental compounds owing to its asymmetric nature. This is because the OM possesses lipopolysaccharides (LPSs) in the outer leaflet and phospholipids (PLs) in the inner leaflet. The maintenance of lipid asymmetry (Mla) system is involved in preserving the distribution of PLs in OM. The periplasmic component of the system MlaC serves as the substrate-binding protein (SBP) that shuttles PLs between the inner and outer membranes. However, an in-depth report highlighting its mechanism of ligand binding is still lacking. This study reports the crystal structure of MlaC from Escherichia coli (EcMlaC) at a resolution of 2.5 Å in a quasi-open state, complexed with PL. The structural analysis reveals that EcMlaC and orthologs comprise two major domains, viz. nuclear transport factor 2-like (NTF2-like) and phospholipid-binding protein (PBP). Each domain can be further divided into two subdomains arranged in a discontinuous fashion. This study further reveals that EcMlaC is polyspecific in nature and follows a reverse mechanism of the opening of the substrate-binding site during the ligand binding. Furthermore, MlaC can bind two PLs by forming subsites in the binding pocket. These findings, altogether, have led to the proposition of the unique “segmented domain movement” mechanism of PL binding, not reported for any known SBP to date. Further, unlike typical SBPs, MlaC has originated from a cystatin-like fold. Overall, this study establishes MlaC to be a non-canonical SBP with a unique ligand-binding mechanism.

ABC transporter

Periplasmic protein

Membrane lipid asymmetry

Segmented domain movement

Mla transport system

ATPase-associated with diverse cellular activities

lipid transfer protein

multiple sequence alignment

nucleotide-binding domain

maintenance of lipid asymmetry

nuclear transport factor 2

phosphatidylethanolamine

phospholipid-binding protein

root-mean-square deviation

substrate-binding protein

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