High-Throughput Mutagenesis and Cross-Complementation Experiments Reveal Substrate Preference and Critical Residues of the Capsule Transporters in <i>Streptococcus pneumoniae</i>
Wan-Zhen Chua, Matthias Maiwald, Kean Lee Chew, Raymond Tzer-Pin Lin, Sanduo Zheng, Lok-To Sham
Abstract
All licensed pneumococcal vaccines target the capsular polysaccharide (CPS). This layer is highly variable and is important for virulence in many bacterial pathogens. Most of the CPSs are produced by the Wzx/Wzy mechanism. In this pathway, CPS repeating units are synthesized in the cytoplasm, which must be flipped across the cytoplasmic membrane before polymerization. This step is mediated by the widely conserved MOP (Multidrug/Oligosaccharidyl-lipid/Polysaccharide) family transporters. Here, we systematically evaluated the interchangeability of these transporters and identified the residues important for substrate specificity and function. Understanding how CPS is synthesized will inform glycoengineering, vaccine development, and antimicrobial discovery.