AI Article Synopsis
- A new red fluorescent protein, mBeRFP, shows sensitivity to chloride levels and has been modified to enhance this sensitivity significantly.
- Through targeted genetic changes (S94V and R205Y), researchers increased chloride affinity to around 106 mM at typical physiological pH.
- The protein's chromophore undergoes isomerization, leading to two emission bands with varying chloride sensitivities, enabling a new method to measure chloride concentrations inside cells.
Article Abstract
A new chloride-sensitive red fluorescent protein derived from is described. We found that mBeRFP exhibited moderate sensitivity to chloride and, via site-directed mutagenesis (S94V and R205Y), we increased the chloride affinity by more than an order of magnitude ( = 106 ± 6 mM) at physiological pH. In addition, isomerization of the chromophore produces a dual emission band with different chloride sensitivities, which allowed us to develop a ratiometric methodology to measure intracellular chloride concentrations.
Download full-text PDF |
Source |
|---|---|
| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8478333 | PMC |
| http://dx.doi.org/10.1021/acssensors.1c00094 | DOI Listing |
Publication Analysis
Top Keywords
Similar Publications
A Prototype of Ultrasensitive Time-Resolved Fluoroimmunoassay with Enhanced Fluorescence System for the Trace Determination of Urinary 8-Hydroxy-2`-Deoxyguanosine, the DNA Oxidative Stress Biomarker.
J Fluoresc
January 2025
Department of Chemistry, Faculty of Science, Chulalongkorn University, Bangkok, Phayathai Road Pathumwan, 10330, Thailand.
This study presents a new highly sensitive and specific time-resolved fluoroimmunoassay (TRFIA) for the measurement of trace amounts of the urinary 8-hydroxy-2`-deoxyguanosine (8-OHdG) which is a biomarker for oxidative stress on DNA. The assay relied on a competitive binding approach and a mouse monoclonal antibody which recognized 8-OHdG with high specificity. In this assay, 8-OHdG conjugated with bovine serum albumin protein (8-OHdG-BSA) was employed as a solid phase antigen.
View Article and Find Full Text PDFBimolecular Fluorescence Complementation (BiFC) Technique for Exocytic Proteins in Murine Hippocampal Neurons.
Methods Mol Biol
January 2025
Institute of Neurophysiology and NeuroCure Cluster of Excellence, Charité Universitätsmedizin Berlin, Berlin, Germany.
The bimolecular fluorescence complementation (BiFC) technique is a powerful tool for visualizing protein-protein interactions in vivo. It involves genetically fused nonfluorescent fragments of green fluorescent protein (GFP) or its variants to the target proteins of interest. When these proteins interact, the GFP fragments come together, resulting in the reconstitution of a functional fluorescent protein complex that can be observed using fluorescence microscopy.
View Article and Find Full Text PDFA Dual-Color Fluorescence-Based Ratiometric Assay to Measure Endocytic Trafficking of Surface Proteins.
Methods Mol Biol
January 2025
Department of Pharmacology, University of Michigan Medical School, Ann Arbor, MI, USA.
Many membrane proteins on the cell surface are constantly internalized from, and re-delivered to, the plasma membrane. This endocytic cycling, which relies on accurate SNARE-mediated fusion of vesicles containing cargo proteins, is highly important for the function of many proteins such as signaling receptors. While the SNARE proteins that mediate fusion during specific events, such as neurotransmitter and hormone release, in mammalian cells has been heavily studied, the SNARE proteins that mediate surface delivery of specific cargo such as the receptors for these released factors are still not known.
View Article and Find Full Text PDFMechanistic Insights into Synaptotagmin-1 Mediated Membrane Fusion and Interactions.
Methods Mol Biol
January 2025
Institute of Physical Chemistry, University of Göttingen, Göttingen, Germany.
We present two innovative approaches to investigate the dynamics of membrane fusion and the strength of protein-membrane interactions. The first approach employs pore-spanning membranes (PSMs), which allow for the observation of protein-assisted fusion processes. The second approach utilizes colloidal probe microscopy with membrane-coated probes with reconstituted proteins.
View Article and Find Full Text PDFStructural Dynamics of SNARE Complex Assembly in the Ribbon Synapses Observed by smFRET.
Methods Mol Biol
January 2025
Quantum-Si, Guilford, CT, USA.
Single-molecule fluorescence resonance energy transfer (smFRET) is a powerful technique for studying the structural dynamics of protein molecules or detecting interactions between protein molecules in real time. Due to the high sensitivity in spatial and temporal resolution, smFRET can decipher sub-populations within heterogeneous native state conformations, which are generally lost in traditional measurements due to ensemble averaging. In addition, the single-molecule reconstitution allows protein molecules to be observed for an extensive period of time and can recapitulate the geometry of the cellular environment to retain biological function.
View Article and Find Full Text PDFWant AI Summaries of new PubMed Abstracts delivered to your In-box?
Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!