Person:

Liang, Feng

Fluorescent labeling techniques have been widely used in live cell studies; however, the labeling processes can be laborious and challenging for use in non-transfectable cells, and labels can interfere with protein functions. While label-free biosensors have been realized by nanofabrication, a method to track intracellular protein dynamics in real-time, in situ and in living cells has not been found. Here we present the first demonstration of label-free detection of intracellular p53 protein dynamics through a nanoscale surface plasmon-polariton fiber-tip-probe (FTP).

Cation-chloride cotransporters (CCCs) mediate the electroneutral transport of chloride, potassium, and/or sodium across the membrane. They play critical roles in regulating cell volume, controlling ion absorption and secretion across epithelia, and maintaining intracellular chloride homeostasis. These transporters are the primary targets for some of the most commonly prescribed drugs in clinic. Here, we determined the cryo-EM structure of a Na-K-Cl cotransporter NKCC1, an extensively-studied member of the CCC transporters. The structure defines the architecture of this protein family and reveals how cytosolic and transmembrane domains are strategically positioned for communication. Structural analyses, functional characterizations, and computational studies uncover the ion translocation pathway, ion-binding sites, and key residues for transport activity. These results provide insights into ion selectivity, coupling, and translocation, and establish a framework for understanding the physiological functions of CCC transporters and interpreting disease-related mutations.

Current methods to study molecular interactions require labeling the subject molecules with fluorescent reporters. The effect of the fluorescent reporters on molecular dynamics, however, has not been quantified due to lack of alternative methods. We develop a hybrid photonic-plasmonic antenna-in-a-nanovacity single molecule biosensor to study DNA-protein dynamics without using fluorescent labels. Our results indicate that the fluorescein and fluorescent protein labels will decrease the interaction between a single DNA and a protein due to weakened electrostatic interaction. While the study is performed on the DNA-XPA system, the conclusion has a generall implication that the traditional fluorescent labeling methods might be misestimating the molecular interactions.