التفاصيل البيبلوغرافية
| العنوان: |
Chemical Tools to Study Protein Pyrophosphorylation |
| المؤلفون: |
Conway, John Henry |
| المساهمون: |
Fiedler, Dorothea, Chemistry Department |
| بيانات النشر: |
Princeton University |
| سنة النشر: |
2015 |
| المجموعة: |
DataSpace at Princeton University |
| مصطلحات موضوعية: |
Affinity Chromatography, Chemical Biology, Chemical Tools, Phosphate, Pyrophosphorylation, Chemistry |
| الوصف: |
This thesis describes the development of tools to study protein pyrophosphorylation, a recently discovered post-translational modification (PTM). Protein pyrophosphorylation occurs when an inositol pyrophosphate (PP-IP) transfer its ß-phosphoryl group to a phosphorylated serine residue on a protein. Genetic studies in mice and mammals have suggested a role for pyrophosphorylation in numerous biological processes, including signaling and body weight regulation. Detection of this modification in complex samples, however, has been elusive due to a variety of technical challenges. Herein, the development of chemical methods to synthesize model pyrophosphorylated peptides as well an affinity reagent that can selectively enrich for these pyrophosphorylated peptides is described. First, to facilitate the study of protein pyrophosphorylation, access to model pyrophosphopeptides was needed. Three solution phase methods to pyrophosphorylate phosphoserine, which utilize electrophilic phosphorus(III) and phosphorus(V) reagents, were explored. The optimized pyrophosphorylation methodology proved to be robust, highly selective, and compatible with complex substrates as well as aqueous solvent. Second, an affinity chromatography reagent was designed to enable enrichment of natively pyrophosphorylated species. The initial design of the reagent was based on existing zinc(II)-dipicolylamine (Zn-dpa) complexes that had been shown to bind inorganic pyrophosphate (PPi) with high specificity in aqueous solution. The affinity of these complexes toward diphosphate esters and other biologically relevant anions was characterized using isothermal titration calorimetry (ITC). Once a complex with ideal affinity and selectivity toward diphosphate esters was identified, a solid-phase analog was synthesized, which generated a pyrophosphopeptide affinity reagent. The reagent was shown to be easy to synthesize, recyclable, and compatible with chemically mild conditions. This affinity reagent, in conjunction with mass spectrometry (MS) methods ... |
| نوع الوثيقة: |
doctoral or postdoctoral thesis |
| اللغة: |
English |
| Relation: |
The Mudd Manuscript Library retains one bound copy of each dissertation. Search for these copies in the library's main catalog; http://arks.princeton.edu/ark:/88435/dsp01rn3013702 |
| الاتاحة: |
http://arks.princeton.edu/ark:/88435/dsp01rn3013702 |
| رقم الانضمام: |
edsbas.974F4D32 |
| قاعدة البيانات: |
BASE |