Identification of a Lipoteichoic Acid Glycosyltransferase Enzyme Reveals that GW-Domain-Containing Proteins Can Be Retained in the Cell Wall of Listeria monocytogenes in the Absence of Lipoteichoic Acid or Its Modifications.

Matthew G Percy,Angelika Gründling,Eleni Karinou,Alexander J Webb

doi:10.1128/jb.00116-16

Abstract

ABSTRACTListeria monocytogenes is a foodborne Gram-positive bacterial pathogen, and many of its virulence factors are either secreted proteins or proteins covalently or noncovalently attached to the cell wall. Previous work has indicated that noncovalently attached proteins with GW (glycine-tryptophan) domains are retained in the cell wall by binding to the cell wall polymer lipoteichoic acid (LTA). LTA is a glycerol phosphate polymer, which is modified in L. monocytogenes with galactose and d-alanine residues. We identified Lmo0933 as the cytoplasmic glycosyltransferase required for the LTA glycosylation process and renamed the protein GtlA, for glycosyltransferase LTA A. Using L. monocytogenes mutants lacking galactose or d-alanine modifications or the complete LTA polymer, we show that GW domain proteins are retained within the cell wall, indicating that other cell wall polymers are involved in the retention of GW domain proteins. Further experiments revealed peptidoglycan as the binding receptor as a purified GW domain fusion protein can bind to L. monocytogenes cells lacking wall teichoic acid (WTA) as well as purified peptidoglycan derived from a wild-type or WTA-negative strain. With this, we not only identify the first enzyme involved in the LTA glycosylation process, but we also provide new insight into the binding mechanism of noncovalently attached cell wall proteins. IMPORTANCE Over the past 20 years, a large number of bacterial genome sequences have become available. Computational approaches are used for the genome annotation and identification of genes and encoded proteins. However, the function of many proteins is still unknown and often cannot be predicted bioinformatically. Here, we show that the previously uncharacterized Listeria monocytogenes gene lmo0933 likely codes for a glycosyltransferase required for the decoration of the cell wall polymer lipoteichoic acid (LTA) with galactose residues. Using L. monocytogenes mutants lacking LTA modifications or the complete polymer, we show that specific cell wall proteins, often associated with virulence, are retained within the cell wall, indicating that additional cell wall polymers are involved in their retention.

Highlights

Over the past 20 years, a large number of bacterial genome sequences have become available
teichoic acids (TAs) are further grouped into wall teichoic acid (WTA), a polymer covalently linked to the peptidoglycan layer, and lipoteichoic acid (LTA), a polymer anchored to the outer leaflet of the cytoplasmic membrane by a glycolipid anchor [15, 16]
Very little is known about the mechanism by which type I LTA is glycosylated, and to the best of our knowledge, we have identified in the present study the first genetic determinant required for this process, namely, the L. monocytogenes gene lmo0933 (Fig. 2 and 3)

Summary

Introduction

Over the past 20 years, a large number of bacterial genome sequences have become available. We show that the previously uncharacterized Listeria monocytogenes gene lmo0933 likely codes for a glycosyltransferase required for the decoration of the cell wall polymer lipoteichoic acid (LTA) with galactose residues. Bound proteins contain specific cell wall binding domains, and in the case of L. monocytogenes, these are GW (glycine-tryptophan) modules or WXL or LysM domains (see reviews in references 6 and 7). The cell wall of L. monocytogenes is typical for a Gram-positive bacterium belonging to the phylum Firmicutes It is composed of a thick peptidoglycan layer and teichoic acids (TAs). TAs are further grouped into wall teichoic acid (WTA), a polymer covalently linked to the peptidoglycan layer, and lipoteichoic acid (LTA), a polymer anchored to the outer leaflet of the cytoplasmic membrane by a glycolipid anchor [15, 16]. The proteins encoded in the dltAdltB-dltC-dltD (dltA-D) operon are responsible for the addition of the D-alanine modifications on LTA [29]; the enzymes responsible for the modification of LTA with galactose residues have not yet been identified

Methods

Results

Conclusion