Expression and characterization of a Mycoplasma genitalium glycosyltransferase in membrane glycolipid biosynthesis: Potential target against Mycoplasma infections

Eduardo Andrés, Núria Martínez, Antoni Planas

Producció científica: Article en revista indexadaArticleAvaluat per experts

28 Cites (Scopus)

Resum

Mycoplasmas contain glycoglycerolipids in their plasma membrane as key structural components involved in bilayer properties and stability. A membrane-associated glycosyltransferase (GT), GT MG517, has been identified in Mycoplasma genitalium, which sequentially produces monoglycosyl- and diglycosyldiacylglycerols. When recombinantly expressed in Escherichia coli, the enzyme was functional in vivo and yielded membrane glycolipids from which Glcβ1,6GlcβDAG was identified as the main product. A chaperone co-expression system and extraction with CHAPS detergent afforded soluble protein that was purified by affinity chromatography. GT MG517 transfers glucosyl and galactosyl residues from UDP-Glc and UDPGal to dioleoylglycerol (DOG) acceptor to form the corresponding β-glycosyl-DOG, which then acts as acceptor to give β-diglycosyl-DOG products. The enzyme (GT2 family) follows Michaelis-Menten kinetics. k cat is about 5-fold higher for UDP-Gal with either DOG or monoglucosyldioleoylglycerol acceptors, but it shows better binding for UDP-Glc than UDP-Gal, as reflected by the lower K m, which results in similar k cat/K m values for both donors. Although sequentially adding glycosyl residues with β-1,6 connectivity, the first glycosyltransferase activity (to DOG) is about 1 order of magnitude higher than the second (to monoglucosyldioleoylglycerol). Because the ratio between the non-bilayer-forming monoglycosyldiacylglycerols and the bilayer-prone diglycosyldiacylglycerols contributes to regulate the properties of the plasma membrane, both synthase activities are probably regulated. Dioleoylphosphatidylglycerol (anionic phospholipid) activates the enzyme, k cat linearly increasing with dioleoylphosphatidylglycerol concentration. GT MG517 is shown to be encoded by an essential gene, and the addition of GT inhibitors results in cell growth inhibition. It is proposed that glycolipid synthases are potential targets for drug discovery against infections by mycoplasmas.

Idioma originalAnglès
Pàgines (de-a)35367-35379
Nombre de pàgines13
RevistaJournal of Biological Chemistry
Volum286
Número41
DOIs
Estat de la publicacióPublicada - 14 d’oct. 2011

Fingerprint

Navegar pels temes de recerca de 'Expression and characterization of a Mycoplasma genitalium glycosyltransferase in membrane glycolipid biosynthesis: Potential target against Mycoplasma infections'. Junts formen un fingerprint únic.

Com citar-ho