Abstract
UTP-glucose-1-phosphate uridylyltransferases are enzymes that produce UDP-glucose from UTP and glucose-1-phosphate. In Bacillus subtilis 168, UDP-glucose is required for the decoration of wall teichoic acid (WTA) with glucose residues and the formation of glucolipids. The B. subtilis UGPase GtaB is essential for UDP-glucose production under standard aerobic growth conditions, and gtaB mutants display severe growth and morphological defects. However, bioinformatics predictions indicate that two other UTP-glucose-1-phosphate uridylyltransferases are present in B. subtilis. Here, we investigated the function of one of them named YngB. The crystal structure of YngB revealed that the protein has the typical fold and all necessary active site features of a functional UGPase. Furthermore, UGPase activity could be demonstrated in vitro using UTP and glucose-1-phosphate as substrates. Expression of YngB from a synthetic promoter in a B. subtilis gtaB mutant resulted in the reintroduction of glucose residues on WTA and production of glycolipids, demonstrating that the enzyme can function as UGPase in vivo. When WT and mutant B. subtilis strains were grown under anaerobic conditions, YngB-dependent glycolipid production and glucose decorations on WTA could be detected, revealing that YngB is expressed from its native promoter under anaerobic condition. Based on these findings, along with the structure of the operon containing yngB and the transcription factor thought to be required for its expression, we propose that besides WTA, potentially other cell wall components might be decorated with glucose residues during oxygen-limited growth condition.
Highlights
The cell envelope of bacteria is composed of several sugarcontaining polymers, including peptidoglycan, capsular polysaccharides, and lipopolysaccharide, in gram-negative bacteria
Lipoteichoic acid (LTA) is a polyglycerol phosphate polymer that is linked by the glycolipid anchor diglucosyl-diacylglycerol (Glc2-DAG) to the outside of the bacterial membrane and further decorated with D-alanine and GlcNAc residues, and wall teichoic acid (WTA) is a polyglycerol phosphate polymer covalently linked to peptidoglycan and decorated with Dalanine and glucose residues [9, 10]
GtaB has been characterized as a UTP-glucose-1-phosphate uridylyltransferase (UGPase), and in its absence, B. subtilis lacks glucose decorations on WTA and is unable to produce glycolipids during vegetative growth [28, 31]
Summary
The cell envelope of bacteria is composed of several sugarcontaining polymers, including peptidoglycan, capsular polysaccharides, and lipopolysaccharide, in gram-negative bacteria. Lipoteichoic acid (LTA) is a polyglycerol phosphate polymer that is linked by the glycolipid anchor diglucosyl-diacylglycerol (Glc2-DAG) to the outside of the bacterial membrane and further decorated with D-alanine and GlcNAc residues, and wall teichoic acid (WTA) is a polyglycerol phosphate polymer covalently linked to peptidoglycan and decorated with Dalanine and glucose residues [9, 10]. Both polymers are made up of glycerol phosphate repeating units but are produced by separate pathways. Apart from GtaB, two other predicted UGPases, YtdA and YngB, are encoded in the B. subtilis 168 genome: as part of this study, we investigated the function of YngB [35, 36]
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