Structural Features of Glycosyltransferases Synthesizing Major Bilayer and Nonbilayer-prone Membrane Lipids inAcholeplasma laidlawii and Streptococcus pneumoniae

Abstract

In membranes of Acholeplasma laidlawii two consecutively acting glucosyltransferases, the (i) ␣-monoglucosyldiacylglycerol (MGlcDAG) synthase (alMGS) (EC 2.4.1.157) and the (ii) ␣-diglucosyl-DAG (DGlcDAG) synthase (alDGS) (EC 2.4.1.208), are involved in maintaining (i) a certain anionic lipid surface charge density and (ii) constant nonbilayer/bilayer conditions (curvature packing stress), respectively. Cloning of the alDGS gene revealed related uncharacterized sequence analogs especially in several Gram-positive pathogens, thermophiles and archaea, where the encoded enzyme function of a potential Streptococcus pneumoniae DGS gene (cpoA) was verified. A strong stimulation of alDGS by phosphatidylglycerol (PG), cardiolipin, or nonbilayer-prone 1,3-DAG was observed, while only PG stimulated CpoA. Several secondary structure prediction and fold recognition methods were used together with SWISS-MODEL to build three-dimensional model structures for three MGS and two DGS lipid glycosyltransferases. Two Escherichia coli proteins with known structures were identified as the best templates, the membrane surface-associated two-domain glycosyltransferase MurG and the soluble GlcNAc epimerase. Differences in electrostatic surface potential between the different models and their individual domains suggest that electrostatic interactions play a role for the association to membranes. Further support for this was obtained when hybrids of the N-and C-domain, and full size alMGS with green fluorescent protein were localized to different regions of the E. coli inner membrane and cytoplasm in vivo. In conclusion, it is proposed that the varying abilities to bind, and sense lipid charge and curvature stress, are governed by typical differences in charge (pI values), amphiphilicity, and hydrophobicity for the N-and (catalytic) C-domains of these structurally similar membrane-associated enzymes.

Comparative Genomic Analysis Revealed a Gene for Monoglucosyldiacylglycerol Synthase, an Enzyme for Photosynthetic Membrane Lipid Synthesis in Cyanobacteria

“…This protein has higher similarity to the plant-type MGDG synthase than to cyanobacterial MGlcDG synthase even though the protein transfers Glc to acceptors. However, by using cyanobacteria, it was shown that the enzyme catalyzes transfer of Glc to MGDG rather than to DAG in vivo, similar to diglucosyldiacylglycerol synthase of A. laidlawii (Edman et al, 2003). Thus, the enzyme is a diglycosyldiacylglycerol synthase, probably not an MGlcDG synthase.…”

Functional differences between galactolipids and glucolipids revealed in photosynthesis of higher plants

“…1). For this purpose, glycosyltransferases were selected from bacterial origin that had previously been characterized by heterologous expression in prokaryotic or eukaryotic hosts (Escherichia coli, Pichia pastoris, Saccharomyces cerevisiae, and Synechococcus) and subsequent analysis of newly formed glycolipids (20)(21)(22)(23)(24)(25)(26). Additional putative glycosyltransferases were identified in bacteria based on sequence similarity to known genes (Table 1) (26), and by this 20 bacterial genes were selected for expression in plants.…”

A Bifunctional Glycosyltransferase from Agrobacterium tumefaciens Synthesizes Monoglucosyl and Glucuronosyl Diacylglycerol under Phosphate Deprivation

“…The best studied glycosyltransferases include the MGD/MGlcD and DGD synthases from plants and cyanobacteria (15)(16)(17)(18), the MGlcD and DGlcD synthases from Acholeplasma (7,19), and the processive glycosyltransferases from the Gram-positive bacteria Staphylococcus or Bacillus (20,21). There are only a few glycosyltransferases characterized as promiscuous enzymes, such as Pgt from Agrobacterium or Mesorhizobium, or the processive glycosyltransferases from the human pathogens Mycoplasma pneumoniae and Mycoplasma genitalum using UDP-Glc and UDP-galactose (UDP-Gal) as sugar donors (11,22,23).…”