Real-Time Insights into Biological Events: In-Cell Processes and Protein-Ligand Interactions

Linda Cerofolini; Stefano Giuntini; Letizia Barbieri; Matteo Pennestri; Anna Codina; Marco Fragai; Lucia Banci; Enrico Luchinat; Enrico Ravera

doi:10.1016/j.bpj.2018.11.3132

Real-Time Insights into Biological Events: In-Cell Processes and Protein-Ligand Interactions

Biophys J. 2019 Jan 22;116(2):239-247. doi: 10.1016/j.bpj.2018.11.3132. Epub 2018 Dec 6.

Authors

Linda Cerofolini¹, Stefano Giuntini², Letizia Barbieri¹, Matteo Pennestri³, Anna Codina³, Marco Fragai², Lucia Banci², Enrico Luchinat⁴, Enrico Ravera⁵

Affiliations

¹ Magnetic Resonance Center, University of Florence and Consorzio Interuniversitario Risonanze Magnetiche di Metalloproteine, Sesto Fiorentino, Italy.
² Magnetic Resonance Center, University of Florence and Consorzio Interuniversitario Risonanze Magnetiche di Metalloproteine, Sesto Fiorentino, Italy; Department of Chemistry, Ugo Schiff, University of Florence, Sesto Fiorentino, Italy.
³ Bruker UK Limited, Banner Lane, Coventry, United Kingdom.
⁴ Magnetic Resonance Center, University of Florence and Consorzio Interuniversitario Risonanze Magnetiche di Metalloproteine, Sesto Fiorentino, Italy; Department of Experimental and Clinical Biomedical Sciences, Mario Serio, University of Florence, Florence, Italy. Electronic address: [email protected].
⁵ Magnetic Resonance Center, University of Florence and Consorzio Interuniversitario Risonanze Magnetiche di Metalloproteine, Sesto Fiorentino, Italy; Department of Chemistry, Ugo Schiff, University of Florence, Sesto Fiorentino, Italy. Electronic address: [email protected].

Abstract

FlowNMR has the aim of continuously monitoring processes that occur in conditions that are not compatible with being carried out within a closed tube. However, it is sample intensive and not suitable for samples, such as proteins or living cells, that are often available in limited volumes and possibly low concentrations. We here propose a dialysis-based modification of a commercial flowNMR setup that allows for recycling the medium while confining the sample (proteins and cells) within the active volume of the tube. This approach is demonstrated in the specific cases of in-cell NMR and protein-based ligand studies.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Batch Cell Culture Techniques / instrumentation
Batch Cell Culture Techniques / methods*
Bioreactors*
Carbonic Anhydrase II / antagonists & inhibitors
Carbonic Anhydrase II / metabolism
Drug Discovery / methods
Enzyme Inhibitors / pharmacology
HEK293 Cells
Humans
Magnetic Resonance Spectroscopy / instrumentation
Magnetic Resonance Spectroscopy / methods*
Protein Binding
Superoxide Dismutase / antagonists & inhibitors
Superoxide Dismutase / metabolism

Substances

Enzyme Inhibitors
Superoxide Dismutase
Carbonic Anhydrase II