دورية أكاديمية
Cell-Free Protein Translation System for Expression of Lipid-Binding Proteins Tagged with Small epitopes and Their Use in Protein-Lipid Overlay Assays.
| العنوان: | Cell-Free Protein Translation System for Expression of Lipid-Binding Proteins Tagged with Small epitopes and Their Use in Protein-Lipid Overlay Assays. |
|---|---|
| المؤلفون: | Paez-Valencia J; Laboratory of Cell and Molecular Biology, University of Wisconsin-Madison, Madison, WI, USA. paezvalencia@wisc.edu., Otegui MS; Department of Botany, University of Wisconsin-Madison, Madison, WI, USA.; Department of Botany and Laboratory of Cell and Molecular Biology, University of Wisconsin-Madison, Madison, WI, USA. |
| المصدر: | Methods in molecular biology (Clifton, N.J.) [Methods Mol Biol] 2020; Vol. 2177, pp. 143-152. |
| نوع المنشور: | Journal Article; Research Support, U.S. Gov't, Non-P.H.S. |
| اللغة: | English |
| بيانات الدورية: | Publisher: Humana Press Country of Publication: United States NLM ID: 9214969 Publication Model: Print Cited Medium: Internet ISSN: 1940-6029 (Electronic) Linking ISSN: 10643745 NLM ISO Abbreviation: Methods Mol Biol Subsets: MEDLINE |
| أسماء مطبوعة: | Publication: Totowa, NJ : Humana Press Original Publication: Clifton, N.J. : Humana Press, |
| مواضيع طبية MeSH: | Cell-Free System/*metabolism , Epitopes/*chemistry , Lipid-Linked Proteins/*metabolism, Adaptor Proteins, Signal Transducing/chemistry ; Adaptor Proteins, Signal Transducing/metabolism ; Lipid-Linked Proteins/chemistry ; Luminescence ; Phosphatidylinositols/metabolism |
| مستخلص: | We adapted an efficient cell-free protein synthesis-based protocol for the production of lipid-binding proteins. The experimental procedures are based on the following steps: (1) cell-free synthesis of soluble, lipid-binding proteins fused to small tags; (2) analysis by dot blot of the accessibility of antibodies to the small tags. (3) protein lipid overlay assay with, immunodetection of bound protein by either chemiluminescence or fluorescence. We also provide a fast and inexpensive protocol for homemade lipid nitrocellulose strips spotted with acidic lipids (mostly phosphoinositides) extracted from plant tissues. These homemade lipid strips can be used for preliminary screen and characterization of putative phosphoinositide-binding proteins. |
| فهرسة مساهمة: | Keywords: Cell free protein synthesis system; Phosphoinositides; Protein–lipid overlay assay; Small tags |
| المشرفين على المادة: | 0 (Adaptor Proteins, Signal Transducing) 0 (Epitopes) 0 (Lipid-Linked Proteins) 0 (Phosphatidylinositols) 0 (Tub protein, mouse) |
| تواريخ الأحداث: | Date Created: 20200708 Date Completed: 20210318 Latest Revision: 20210318 |
| رمز التحديث: | 20240628 |
| DOI: | 10.1007/978-1-0716-0767-1_12 |
| PMID: | 32632811 |
| قاعدة البيانات: | MEDLINE |
| تدمد: | 1940-6029 |
|---|---|
| DOI: | 10.1007/978-1-0716-0767-1_12 |