Extended Data Fig. 8: Effect of nucleotide binding mutants of MlaF on ATPase activity and in vitro PL transport.
From: Structural insights into outer membrane asymmetry maintenance in Gram-negative bacteria by MlaFEDB
a, The size-exclusion chromatogram of purified MlaFEDB complexes with MlaF single catalytic mutants of F16A, R18A, K47A, E170A and H203A. The mutants were eluted almost at the same time as the wild-type MlaFEDB, indicating that the mutants have the intact structure as the wild-type. b, The relative ATPase activities of MlaFEDB wild-type and mutants in detergent and liposomes. c, SDS-PAGE analysis of purified wildtype and the mutants. d, FRET scan of PL transport using MlaFEDB mutant IM proteoliposomes containing single mutation on MlaF K47A, E170A or H203A, wildtype OM proteoliposomes and MlaC for retrograde direction. The catalytic and ATP binding residue mutants abolished retrograde transport of PLs. e, Amino acid sequence alignment of MlaF and LptB. MlaF and LptB have 24.54% amino acid identity. The C-terminal tail of MlaF is longer than that of LptB. f, Dimeric MlaF is superimposed into the dimeric LptB. MlaF has a long C-terminal tail that interacts with the other MlaF molecule, but LptB does not. Data in a, c and d are representative results from n = 3 independent experiments. Data in b represents mean ± s.d. (n = 3 independent experiments). An uncropped image for panel c is available online. Source data for panels b and d are available in Supplementary Data Set 1.
