Department of Molecular Microbiology, Washington University School of Medicine in St. Louis, St. Louis, MO, USA.
VaxNewMo LLC, St. Louis, MO, USA.
Glycobiology. 2018 Jul 1;28(7):522-533. doi: 10.1093/glycob/cwy037.
Acinetobacter baumannii is an opportunistic human pathogen with the highest reported rates of multidrug resistance among Gram-negative pathogens. The capsular polysaccharide of A. baumannii is considered one of its most significant virulence factors providing resistance against complemented-mediated killing. Capsule synthesis in A. baumannii is usually initiated by the phosphoglycosyltransferase PglC. PglC transfers a phosphosugar from a nucleotide diphosphate-sugar to a polyprenol phosphate generating a polyprenol diphosphate-linked monosaccharide. Traditionally, PglC was thought to have stringent specificity towards UDP-N-N'-diacetylbacillosamine (UDP-diNAcBac). In this work we demonstrate that A. baumannii PglC has the ability to utilize three different UDP-sugar substrates: UDP-N-acetylglucosamine (UDP-GlcNAc), UDP-N-acetylgalactosamine (UDP-GalNAc) or UDP-diNAcBac. Using phylogenetic analyses, we first demonstrate that A. baumannii PglC orthologs separate into three distinct clades. Moreover, all members within a clade are predicted to have the same preference for one of the three possible sugar substrates. To experimentally determine the substrate specificity of each clade, we utilized in vivo complementation models and NMR analysis. We demonstrate that UDP-diNAcBac is accommodated by all PglC orthologs, but some orthologs evolved to utilize UDP-GlcNAc or UDP-GalNAc in a clade-dependent manner. Furthermore, we show that a single point mutation can modify the sugar specificity of a PglC ortholog specific for UDP-diNAcBac and that introduction of a non-native PglC ortholog into A. baumannii can generate a new capsule serotype. Collectively, these studies begin to explain why A. baumannii strains have such highly diverse glycan repertoires.
鲍曼不动杆菌是一种机会性人类病原体，其对革兰氏阴性病原体的耐药率是报告中最高的。鲍曼不动杆菌的荚膜多糖被认为是其最重要的毒力因子之一，可提供对补体介导杀伤的抗性。鲍曼不动杆菌的荚膜合成通常由磷酸糖基转移酶 PglC 启动。PglC 将磷酸糖从核苷酸二磷酸糖转移到多萜醇磷酸上，生成多萜醇二磷酸连接的单糖。传统上，PglC 被认为对 UDP-N-N'-二乙酰胞壁酰二胺（UDP-diNAcBac）具有严格的特异性。在这项工作中，我们证明了鲍曼不动杆菌 PglC 能够利用三种不同的 UDP-糖底物：UDP-N-乙酰葡萄糖胺（UDP-GlcNAc）、UDP-N-乙酰半乳糖胺（UDP-GalNAc）或 UDP-diNAcBac。通过系统发育分析，我们首先证明鲍曼不动杆菌 PglC 同源物分为三个不同的分支。此外，每个分支内的所有成员都预测对三种可能的糖底物中的一种具有相同的偏好。为了实验确定每个分支的底物特异性，我们利用体内互补模型和 NMR 分析。我们证明所有 PglC 同源物都能容纳 UDP-diNAcBac，但有些同源物以依赖于分支的方式进化为利用 UDP-GlcNAc 或 UDP-GalNAc。此外，我们表明单个点突变可以改变特定于 UDP-diNAcBac 的 PglC 同源物的糖特异性，并且将非天然 PglC 同源物引入鲍曼不动杆菌可以产生新的荚膜血清型。总之，这些研究开始解释为什么鲍曼不动杆菌菌株具有如此多样化的聚糖库。
