Direct Evaluation of Live Uropathogenic Escherichia coli Adhesion and Efficiency of Antiadhesive Compounds Using a Simple Microarray Approach (Analytical Chemistry 2018, 90:12314-12321)

報告日期: 2019/11/12
報告時間: 16:00/16:50
報告學生: 陳宥佐
講評老師: 鄧景浩
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Direct Evaluation of Live Uropathogenic Escherichia coli Adhesion and Efficiency of Antiadhesive Compounds Using a Simple Microarray Approach

Ioanna Kalograiaki, Marta Abellán-Flos, Luis Ángel Fernández, Margarita Menéndez, Stéphane P. Vincent, and Dolores Solís

Speaker:You-Zuo Chen (陳宥佐)                              Date/Time:2019/11/12, 16:00-16:50

Commentator:Ching-Hao Teng (鄧景浩老師)             Place:Room 602

Abstract:

Host cell surface glycans are recognized docking hubs which act as first contact point for attachment and entry. Fimbriae of uropathogenic Escherichia coli (UPEC) causing urinary tract infection(UTI) can mediate adhesion by a type 1 fimbrin FimH adhesion protein, which can target highly mannosylated glycoprotein uroplakin 1a on host cell surface[1]. Therefore, blocking this critical FimH-mannose interaction could be a possible UTI treatment strategy. Here, the mannan-poly-L-lysine conjugates were immobilized onto N-hydroxysuccinimide (NHS) modified glass slide resulting in fabrication of mannan microarray. In addition, the feasibility of the new strategy was analyzed by live UTI89 directly probing on mannan microarray. This approach for assessing the bacterial adhesion and efficiency of anti-adhesive compounds targeting is unique. This method can avoid incorrect assessment for anti-adhesive compounds which due to only blockage of the FimH-mediated UTI89 attachment. As they use live UTI89 with complete integrity of fimbriae, the total number and spacing on UTI89. Concanavalin A (ConA) is a mannose-specific lectin which was used to identify mannan residue and confirmation of a successful the mannan immobilization onto the glass slide.Further, growth phase dependent binding of UTI89 to the arrays was observed, in contrast to the FimH knock-out UTI89 and MeαMan treatment, proving the usefulness of the platform for detecting differences in FimH expression or binding competition. The efficiency of three different dodecamannosylated fullerenes as FimH-targeted compounds was evaluated by microarray competition assay and isothermal titration calorimetry affinity analysis. The results revealed that the mannofullerenes have higher inhibitory effect than methyl α-D-mannopyranoside. Moreover, the sensitivity of the assay was indicated by significant differences in binding affinity were detected between mannofullerenes as fullerene core graft represent different structure/length of mannose residues. Overall, the approach combines straightforward and time-saving protocol for microarray preparation, bacteria labeling, and binding assay which can be easily tailored to other bacteria bearing carbohydrate-binding adhesins. 

Reference:

  1. Poole, J., et al., Glycointeractions in bacterial pathogenesis. Nature Reviews Microbiology, 2018. 16(7): p. 440-452.