title: Membrane remodeling capacity of a vesicle-inducing glycosyltransferase.
abstract: Intracellular vesicles are abundant in eukaryotic cells but absent in the Gram-negative bacterium Escherichia coli. However, strong overexpression of a monotopic glycolipid-synthesizing enzyme, monoglucosyldiacylglycerol synthase from Acholeplasma laidlawii (alMGS), leads to massive formation of vesicles in the cytoplasm of E. coli. More importantly, alMGS provides a model system for the regulation of membrane properties by membrane-bound enzymes, which is critical for maintaining cellular integrity. Both phenomena depend on how alMGS binds to cell membranes, which is not well understood. Here, we carry out a comprehensive investigation of the membrane binding of alMGS by combining bioinformatics methods with extensive biochemical studies, structural modeling and molecular dynamics simulations. We find that alMGS binds to the membrane in a fairly upright manner, mainly by residues in the N-terminal domain, and in a way that induces local enrichment of anionic lipids and a local curvature deformation. Furthermore, several alMGS variants resulting from substitution of residues in the membrane anchoring segment are still able to generate vesicles, regardless of enzymatic activity. These results clarify earlier theories about the driving forces for vesicle formation, and shed new light on the membrane binding properties and enzymatic mechanism of alMGS and related monotopic GT-B fold glycosyltransferases.
authors: Department of Biochemistry and Biophysics, Center for Biomembrane Research, Stockholm University, Sweden; Laboratory for the Structure and Function of Biological Membranes, Center for Structural Biology and Bioinformatics, Université Libre de Bruxelles, Belgium; Medical Inflammation Research, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, Sweden.
authors: Changrong
authors: C
authors: Ge
authors: Jordi
authors: J
authors: Gómez-Llobregat
authors: Marcin J
authors: MJ
authors: Skwark
authors: Jean-Marie
authors: JM
authors: Ruysschaert
authors: Ake
authors: A
authors: Wieslander
authors: Martin
authors: M
authors: Lindén
mesh_terms: D000128:Acholeplasma laidlawii / Q000201:enzymology; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry / Q000502:physiology*; D001665:Binding Sites; D002462:Cell Membrane / Q000201:enzymology; D022162:Cytoplasmic Vesicles / Q000201:enzymology*; D004926:Escherichia coli / Q000201:enzymology / Q000648:ultrastructure; D016695:Glycosyltransferases / Q000737:chemistry / Q000502:physiology*; D008051:Lipid Bilayers / Q000737:chemistry; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
chemical_list: D001426:Bacterial Proteins; D008051:Lipid Bilayers; D016695:Glycosyltransferases
keywords: glycosyltransferase; lipid-protein interactions; membrane deformation; molecular dynamics; monotopic protein