Effector glycosyltransferases in legionella

Abstract

Legionella causes severe pneumonia in humans. The pathogen produces an array of effectors, which interfere with host cell functions. Among them are the glucosyltransferases Lgt1, Lgt2 and Lgt3 from L. pneumophila. Lgt1 and Lgt2 are produced predominately in the post-exponential phase of bacterial growth, while synthesis of Lgt3 is induced mainly in the lag-phase before intracellular replication of bacteria starts. Lgt glucosyltransferases are structurally similar to clostridial glucosylating toxins. The enzymes use UDP-glucose as a donor substrate and modify eukaryotic elongation factor eEF1A at serine-53. This modification results in inhibition of protein synthesis and death of target cells.In addition to Lgts, Legionella genomes disclose several genes, coding for effector proteins likely to possess glycosyltransferase activities, including SetA (subversion of eukaryotic vesicle trafficking A), which influences vesicular trafficking in the yeast model system and displays tropism for late endosomal/lysosomal compartments of mammalian cells. This review mainly discusses recent results on the structure-function relationship of Lgt glucosyltransferases.

Keywords: Legionella; eEF1A; glycosyltransferase; protein synthesis; virulence factor.

Figure 1

(A) Alignment of partial amino acid sequences of Lgt1, SetA, Lpg1961 from L. pneumophila Philadelphia-1 strain with that of proteins from a clostridial glucosylating toxin family: Toxins A and B from C. difficile, α-toxin from C. novyi, and lethal toxin from C. sordellii. Gene bank accession numbers of the corresponding coding sequences are shown in brackets. Essential amino acids mentioned in the text are highlighted (DXD-motif, GT-A triad). (B) Alignment of partial amino acid sequences of Lgt1 from L. pneumophila Philadelphia-1 with that of putative glycosyltransferases found in translated genomes of L. drancourtii LLAP12 and L. longbeachae D-4968. Identification codes for Lgt1 and putative glycosyltransferases in strain LLAP12 of L. drancourtii and strain D-4968 of L. longbeachae (two products in each strain) are Lpg1368, LDG0102/LDG0103, and LLB0067/LLB3681 respectively. Proteins LLO1578 and LLO1721 found in translated sequenced genome of L. longbeachae NSW150 were 100% identical to LLB0067 and LLB3681 from L. longbeachae D-4968 respectively and are not shown on the figure for simplicity reason. Identical amino acid residues are denoted by asterisks, highly conserved residues by double dots, and modestly conserved residues by dots. The secondary structural elements were deduced from the structure of Lgt1 (pdb 3JSZ). The alignment was prepared using ESPript 2.2 (http://espript.ibcp.fr).

Figure 2

Structural view of yeast elongation factor eEF1A (adapted from pdb 1IJF). Elongation factor eEF1A consists of three main structural parts: domain 1 (G-domain), domain 2, and domain 3 (indicated by numbers). The decapeptide (GKGSFKYAWV), which is a sufficient substrate for glucosylation by Lgt, is shown in red. Serine-53, which is modified by glucosyltransferases Lgt, is shown in yellow. The complexed fragment of eEF1Bα molecule, which is present in the original structure, is omitted.

Figure 3

Cartoon presentation of Lgt1 crystal structure in complex with UDP–glucose and Mg²⁺ (pdb code 3JSZ). (A) The N-terminal domain is depicted in blue, the central domain in gray and the protrusion domain in brown. The central beta sheet is shaded in light blue. UDP–glucose is shown in sticks and Mg²⁺ as a red sphere. The flexible loop region is highlighted in dark red. Aspartic acid residues of the DXD-motif, Trp-520 of the flexible loop, Trp-139 stacking the base are shown in sticks. (B) Magnified view on the catalytic site of Lgt1 as in (A) with intact UDP–glucose (white) and glucose shifted about 1.6 Å after cleavage (dark blue; deduced from pdb 2WZF). Important amino acids are shown as sticks. Trp-139 is stacking the uracil ring of the donor, Asp-230, Arg-233, and Asp-246 are orientating the distal part of glucose as a triade. Asp-248 of the DXD-motif is coordinating the divalent ion (red) in conjunction with three additional water molecules. Trp-520 of the flexible loop is marked in red. (C) Schematic representation of the catalytic site as in (B) with important amino acids marked and highlighted.