Buß, O. ; Jager, S. ; Dold, S.-M. ; Zimmermann, S. ; Hamacher, Kay ; Schmitz, K. ; Rudat, J. (2016)
Statistical Evaluation of HTS Assays for Enzymatic Hydrolysis of β-Keto Esters.
In: PloS one, 11 (1)
Artikel, Bibliographie
Kurzbeschreibung (Abstract)
β-keto esters are used as precursors for the synthesis of β-amino acids, which are building blocks for some classes of pharmaceuticals. Here we describe the comparison of screening procedures for hydrolases to be used for the hydrolysis of β-keto esters, the first step in the preparation of β-amino acids. Two of the tested high throughput screening (HTS) assays depend on coupled enzymatic reactions which detect the alcohol released during ester hydrolysis by luminescence or absorption. The third assay detects the pH shift due to acid formation using an indicator dye. To choose the most efficient approach for screening, we assessed these assays with different statistical methods-namely, the classical Z'-factor, standardized mean difference (SSMD), the Kolmogorov-Smirnov-test, and t-statistics. This revealed that all three assays are suitable for HTS, the pH assay performing best. Based on our data we discuss the explanatory power of different statistical measures. Finally, we successfully employed the pH assay to identify a very fast hydrolase in an enzyme-substrate screening.
| Typ des Eintrags: | Artikel |
|---|---|
| Erschienen: | 2016 |
| Autor(en): | Buß, O. ; Jager, S. ; Dold, S.-M. ; Zimmermann, S. ; Hamacher, Kay ; Schmitz, K. ; Rudat, J. |
| Art des Eintrags: | Bibliographie |
| Titel: | Statistical Evaluation of HTS Assays for Enzymatic Hydrolysis of β-Keto Esters. |
| Sprache: | Englisch |
| Publikationsjahr: | 2016 |
| Titel der Zeitschrift, Zeitung oder Schriftenreihe: | PloS one |
| Jahrgang/Volume einer Zeitschrift: | 11 |
| (Heft-)Nummer: | 1 |
| Kurzbeschreibung (Abstract): | β-keto esters are used as precursors for the synthesis of β-amino acids, which are building blocks for some classes of pharmaceuticals. Here we describe the comparison of screening procedures for hydrolases to be used for the hydrolysis of β-keto esters, the first step in the preparation of β-amino acids. Two of the tested high throughput screening (HTS) assays depend on coupled enzymatic reactions which detect the alcohol released during ester hydrolysis by luminescence or absorption. The third assay detects the pH shift due to acid formation using an indicator dye. To choose the most efficient approach for screening, we assessed these assays with different statistical methods-namely, the classical Z'-factor, standardized mean difference (SSMD), the Kolmogorov-Smirnov-test, and t-statistics. This revealed that all three assays are suitable for HTS, the pH assay performing best. Based on our data we discuss the explanatory power of different statistical measures. Finally, we successfully employed the pH assay to identify a very fast hydrolase in an enzyme-substrate screening. |
| Fachbereich(e)/-gebiet(e): | 10 Fachbereich Biologie 10 Fachbereich Biologie > Computational Biology and Simulation 07 Fachbereich Chemie 07 Fachbereich Chemie > Clemens-Schöpf-Institut > Fachgebiet Biochemie 07 Fachbereich Chemie > Clemens-Schöpf-Institut > Fachgebiet Biochemie > Biologische Chemie |
| Hinterlegungsdatum: | 13 Jan 2016 07:13 |
| Letzte Änderung: | 23 Apr 2020 09:02 |
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