Chandrasekar, Balakumaran: From glycosidase activity profiling to inhibitor discovery in the plant pathogen Pseudomonas syringae. - Bonn, 2018. - Dissertation, Rheinische Friedrich-Wilhelms-Universität Bonn.
Online-Ausgabe in bonndoc: https://nbn-resolving.org/urn:nbn:de:hbz:5n-50839
@phdthesis{handle:20.500.11811/7568,
urn: https://nbn-resolving.org/urn:nbn:de:hbz:5n-50839,
author = {{Balakumaran Chandrasekar}},
title = {From glycosidase activity profiling to inhibitor discovery in the plant pathogen Pseudomonas syringae},
school = {Rheinische Friedrich-Wilhelms-Universität Bonn},
year = 2018,
month = may,

note = {Retaining glycosidases are important in plants for various biological processes. Plant genomes encode for more than 200 retaining glycosidases but the physiological roles and activities of many remain poorly understood. We established glycosidase activity profiling in plants using activity-based probes based on cyclophellitol-aziridine. These probes targeted a diversity of glycosidases belonging to different GH families in leaf extracts of Arabidopsis thaliana and in the cell wall proteome of Nicotiana benthamiana. When applied to plants infected with the bacterial pathogen Pseudomonas syringae, we discovered that the activity of a cell wall associated beta-galactosidase (BGAL) is suppressed during infection. We also introduce convolution ABPP, a novel inhibitor discovery approach that revealed that the suppression of BGAL is caused by a small, heat-stable inhibitor that is produced by the pathogen. Bacterial genetics together with biochemical approaches indicate that trihydroxy piperidine (THP), an imino sugar might be the BGAL inhibitor produced by the pathogen. Trihydroxy piperidine (THP) was detected during infection and in bacterial cultures using GC-MS and HRMS. Pathogen growth assays using the BGAL inhibitor mutants indicate that BGAL inhibitor production is important for the virulence of PtoDC3000, whereas depletion of BGAL using virus-induced gene silencing (VIGS) increased bacterial growth, indicating the discovery of an important immune enzyme that is suppressed by pathogenic bacteria. Furthermore, two candidate PLCPs that might be required for processing the BGAL enzyme in Nicotiana benthamiana were identified using VIGS. In addition, a new tool for functional classification of JJB-labeled glycosidases has been introduced based on product inhibition of glycosidases. Moreover, two interesting observations during my PhD program are also documented. First, increased glycosidases activities observed in the growth medium of a PtoDC3000 mutant is due to the disruption of a gene encoding for an ABC transporter. Second, unexpected inhibitory effects of commonly used antibiotics on plant glycosidases were discovered.},
url = {http://hdl.handle.net/20.500.11811/7568}
}

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