Mechanism of Staphylococcus aureus lipoteichoic acid length regulation, inhibition of wall teichoic acid biosynthesis, and physiology of teichoic acids

Abstract

Bacteria are surrounded by a complex cell envelope that is essential for viability and protection against antibiotics and other external insults. The cell envelope is comprised of a cell membrane, a peptidoglycan cell wall, proteins, and numerous carbohydrate polymers that exhibit incredible structural diversity. As peptidoglycan is essential and highly conserved, peptidoglycan biosynthesis is targeted by many successful antibiotics. However, the continued threat of antibiotic resistance and detrimental effects of broad-spectrum antibiotics on the human microbiome has led to calls for new antibiotic targets. Many Gram-positive bacteria also produce teichoic acids, which are negatively charged glycopolymers that are divided into two classes. Wall teichoic acids (WTAs) are covalently attached to peptidoglycan while lipoteichoic acids (LTAs) remain associated with the cell membrane through a lipid anchor. Given the importance of teichoic acids in cell division, virulence, and resistance to antimicrobial agents, they have been proposed as antibiotic targets. The diversity of teichoic acids between organisms also offers specificity to antibiotics targeting them. However, the complete physiological roles of teichoic acids and the mechanisms by which cell envelope homeostasis is maintained is unclear. In this work, we discovered that the length of LTA polymers is critical for control of Staphylococcus aureus cell size and envelope integrity and identified several compensatory mechanisms that S. aureus employs to correct for defects in LTA synthesis. Furthermore, we found that methicillin-resistant S. aureus strains with abnormally long LTA polymers are re-sensitized to beta-lactam antibiotics. Using in vitro reconstitution of the LTA polymerase, we identified the mechanism by which the length of LTA polymers is determined and proposed a model by which this is achieved on a molecular level. We also elucidated the mechanism of action of an inhibitor of WTA biosynthesis and exploited it to purify lipid-linked WTA precursors directly from cells. In addition to these findings, the tools and methods developed here for biochemical analysis of teichoic acids will assist future efforts seeking to characterize bacterial cell envelopes and develop antibiotics targeting teichoic acid biosynthesis. Together, this work provides a more complete picture of the functions of teichoic acids in Gram-positive bacterial physiology.