Specific ion effects on macromolecular interactions inEscherichia coliextracts
| Autor: | Ciara Kyne, Virginie Gautier, Brian Ruhle, Peter B. Crowley |
|---|---|
| Rok vydání: | 2014 |
| Předmět: |
0303 health sciences
RNase P Chemistry Size-exclusion chromatography RNA Nuclear magnetic resonance spectroscopy 010402 general chemistry medicine.disease_cause 01 natural sciences Biochemistry 0104 chemical sciences Protein–protein interaction 03 medical and health sciences Ionic strength medicine Molecular Biology Escherichia coli 030304 developmental biology Macromolecule |
| Zdroj: | Protein Science. 24:310-318 |
| ISSN: | 0961-8368 |
| DOI: | 10.1002/pro.2615 |
| Popis: | Protein characterization in situ remains a major challenge for protein science. Here, the interactions of ΔTat-GB1 in Escherichia coli cell extracts were investigated by NMR spectroscopy and size exclusion chromatography (SEC). ΔTat-GB1 was found to participate in high molecular weight complexes that remain intact at physiologically-relevant ionic strength. This observation helps to explain why ΔTat-GB1 was not detected by in-cell NMR spectroscopy. Extracts pre-treated with RNase A had a different SEC elution profile indicating that ΔTat-GB1 predominantly interacted with RNA. The roles of biological and laboratory ions in mediating macromolecular interactions were studied. Interestingly, the interactions of ΔTat-GB1 could be disrupted by biologically-relevant multivalent ions. The most effective shielding of interactions occurred in Mg2+-containing buffers. Moreover, a combination of RNA digestion and Mg2+ greatly enhanced the NMR detection of ΔTat-GB1 in cell extracts. |
| Databáze: | OpenAIRE |
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