Browsing by Author "Zhao, Boyang"

Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Enteroaggregative E. coli Adherence to Human Heparan Sulfate Proteoglycans Drives Segment and Host Specific Responses to Infection
    (Public Library of Science, 2020) Rajan, Anubama; Robertson, Matthew J.; Carter, Hannah E.; Poole, Nina M.; Clark, Justin R.; Green, Sabrina I.; Criss, Zachary K.; Zhao, Boyang; Karandikar, Umesh; Xing, Yikun; Margalef-Català, Mar; Jain, Nikhil; Wilson, Reid L.; Bai, Fan; Hyser, Joseph M.; Petrosino, Joseph; Shroyer, Noah F.; Blutt, Sarah E.; Coarfa, Cristian; Song, Xuezheng; Prasad, BV Venkataram; Amieva, Manuel R.; Grande-Allen, Jane; Estes, Mary K.; Okhuysen, Pablo C.; Maresso, Anthony W.; Bioengineering
    Enteroaggregative Escherichia coli (EAEC) is a significant cause of acute and chronic diarrhea, foodborne outbreaks, infections of the immunocompromised, and growth stunting in children in developing nations. There is no vaccine and resistance to antibiotics is rising. Unlike related E. coli pathotypes that are often associated with acute bouts of infection, EAEC is associated with persistent diarrhea and subclinical long-term colonization. Several secreted virulence factors have been associated with EAEC pathogenesis and linked to disease in humans, less certain are the molecular drivers of adherence to the intestinal mucosa. We previously established human intestinal enteroids (HIEs) as a model system to study host-EAEC interactions and aggregative adherence fimbriae A (AafA) as a major driver of EAEC adherence to HIEs. Here, we report a large-scale assessment of the host response to EAEC adherence from all four segments of the intestine across at least three donor lines for five E. coli pathotypes. The data demonstrate that the host response in the duodenum is driven largely by the infecting pathotype, whereas the response in the colon diverges in a patient-specific manner. Major pathways altered in gene expression in each of the four enteroid segments differed dramatically, with responses observed for inflammation, apoptosis and an overwhelming response to different mucin genes. In particular, EAEC both associated with large mucus droplets and specific mucins at the epithelial surface, binding that was ameliorated when mucins were removed, a process dependent on AafA. Pan-screening for glycans for binding to purified AafA identified the human ligand as heparan sulfate proteoglycans (HSPGs). Removal of HSPG abrogated EAEC association with HIEs. These results may mean that the human intestine responds remarkably different to distinct pathobionts that is dependent on the both the individual and intestinal segment in question, and uncover a major role for surface heparan sulfate proteoglycans as tropism-driving factor in adherence and/or colonization.
  • Thumbnail Image
    Item
    Structural basis of the stereoselective formation of the spirooxindole ring in the biosynthesis of citrinadins
    (Springer Nature, 2021) Liu, Zhiwen; Zhao, Fanglong; Zhao, Boyang; Yang, Jie; Ferrara, Joseph; Sankaran, Banumathi; Venkataram Prasad, B.V.; Kundu, Biki Bapi; Phillips, George N.Jr.; Gao, Yang; Hu, Liya; Zhu, Tong; Gao, Xue
    Prenylated indole alkaloids featuring spirooxindole rings possess a 3R or 3S carbon stereocenter, which determines the bioactivities of these compounds. Despite the stereoselective advantages of spirooxindole biosynthesis compared with those of organic synthesis, the biocatalytic mechanism for controlling the 3R or 3S-spirooxindole formation has been elusive. Here, we report an oxygenase/semipinacolase CtdE that specifies the 3S-spirooxindole construction in the biosynthesis of 21R-citrinadin A. High-resolution X-ray crystal structures of CtdE with the substrate and cofactor, together with site-directed mutagenesis and computational studies, illustrate the catalytic mechanisms for the possible β-face epoxidation followed by a regioselective collapse of the epoxide intermediate, which triggers semipinacol rearrangement to form the 3S-spirooxindole. Comparing CtdE with PhqK, which catalyzes the formation of the 3R-spirooxindole, we reveal an evolutionary branch of CtdE in specific 3S spirocyclization. Our study provides deeper insights into the stereoselective catalytic machinery, which is important for the biocatalysis design to synthesize spirooxindole pharmaceuticals.