title: Qualitative analysis of the fluorophosphonate-based chemical probes using the serine hydrolases from mouse liver and poly-3-hydroxybutyrate depolymerase (PhaZ) from Bacillus thuringiensis.
abstract: The serine hydrolase family consists of more than 200 members and is one of the largest enzyme families in the human genome. Although up to 50 % of this family remains unannotated, there are increasing evidences that activities of certain serine hydrolases are associated with diseases like cancer neoplasia, invasiveness, etc. By now, several activity-based chemical probes have been developed and are applied to profile the global activity of serine hydrolases in diverse proteomes. In this study, two fluorophosphonate (FP)-based chemical probes were synthesized. Further examination of their abilities to label and pull down serine hydrolases was conducted. In addition, the poly-3-hydroxybutyrate depolymerase (PhaZ) from Bacillus thuringiensis was demonstrated as an appropriate standard serine hydrolase, which can be applied to measure the labeling ability and pull-down efficiency of FP-based probes. Furthermore, mass spectrometry (MS) was used to identify the serine residue that covalently bonded to the active probes. Finally, these FP-based probes were shown capable of establishing the serine hydrolase profiles in diverse mouse tissues; the serine hydrolases pulled down from mouse liver organ were further identified by MS. In summary, our study provides an adequate method to evaluate the reactivity of FP-based probes targeting serine hydrolases.
authors.affiliation: Department of Life Sciences and Institute of Genome Sciences, National Yang Ming University, Taipei, Taiwan.
authors.forename: Yi-Long
authors.initials: YL
authors.lastname: Huang
authors.forename: Tsai-Wen
authors.initials: TW
authors.lastname: Chung
authors.forename: Chia-Mao
authors.initials: CM
authors.lastname: Chang
authors.forename: Chih-Hau
authors.initials: CH
authors.lastname: Chen
authors.forename: Chen-Chung
authors.initials: CC
authors.lastname: Liao
authors.forename: Yeou-Guang
authors.initials: YG
authors.lastname: Tsay
authors.forename: Gwo-Chyuan
authors.initials: GC
authors.lastname: Shaw
authors.forename: Shwu-Huey
authors.initials: SH
authors.lastname: Liaw
authors.forename: Chung-Ming
authors.initials: CM
authors.lastname: Sun
authors.forename: Chao-Hsiung
authors.initials: CH
authors.lastname: Lin
mesh_terms: D000818:Animals; D001413:Bacillus thuringiensis / Q000201:enzymology*; D015153:Blotting, Western; D002265:Carboxylic Ester Hydrolases / Q000378:metabolism; D002623:Chemistry Techniques, Analytical; D004591:Electrophoresis, Polyacrylamide Gel; D005461:Fluorine / Q000032:analysis* / Q000737:chemistry; D008099:Liver / Q000201:enzymology*; D013058:Mass Spectrometry; D051379:Mice; D015335:Molecular Probes / Q000032:analysis* / Q000138:chemical synthesis / Q000737:chemistry; D063065:Organophosphonates / Q000032:analysis* / Q000138:chemical synthesis / Q000737:chemistry; D057057:Serine Proteases / Q000378:metabolism*
chemical_list: D015335:Molecular Probes; D063065:Organophosphonates; D005461:Fluorine; D002265:Carboxylic Ester Hydrolases; C022417:poly-beta-hydroxybutyrate depolymerase; D057057:Serine Proteases