윤태영 교수

Membrane fusion in eukaryotic cells is thought to be mediated by a highly conserved family of proteins called SNAREs (soluble N-ethyl maleimide sensitive-factor attachment protein receptors). The vesicle-associated v-SNARE engages with its partner t-SNAREs on the target membrane to form a coiled coil that bridges two membranes and facilitates fusion. As demonstrated by recent findings on the hemifusion state, identifying intermediates of membrane fusion can help unveil the underlying fusion mech

In neurons, synaptotagmin 1 (Syt1) is thought to mediate the fusion of synaptic vesicles with the plasma membrane when presynaptic Ca2+ levels rise. However, in vitro reconstitution experiments have failed to recapitulate key characteristics of Ca2+-triggered membrane fusion. Using an in vitro single-vesicle fusion assay, we found that membrane-anchored Syt1 enhanced Ca2+ sensitivity and fusion speed. This stimulatory activity of membrane-anchored Syt1 dropped as the Ca2+ level rose beyond physi

During intracellular membrane trafficking, N-ethylmaleimide-sensitive factor (NSF) and alpha-soluble NSF attachment protein (α-SNAP) disassemble the soluble NSF attachment protein receptor (SNARE) complex for recycling of the SNARE proteins. The molecular mechanism by which NSF disassembles the SNARE complex is largely unknown. Using single-molecule fluorescence spectroscopy and magnetic tweezers, we found that NSF disassembled a single SNARE complex in only one round of adenosine triphosphate (

To understand membrane protein biogenesis, we need to explore folding within a bilayer context. Here, we describe a single-molecule force microscopy technique that monitors the folding of helical membrane proteins in vesicle and bicelle environments. After completely unfolding the protein at high force, we lower the force to initiate folding while transmembrane helices are aligned in a zigzag manner within the bilayer, thereby imposing minimal constraints on folding. We used the approach to char

Submicrometer elasticity of double-stranded DNA (dsDNA) governs nanoscale bending of DNA segments and their interactions with proteins. Single-molecule force spectroscopy, including magnetic tweezers (MTs), is an important tool for studying DNA mechanics. However, its application to short DNAs under 1 μm is limited. We developed an MT-based method for precise force-extension measurements in the 100-nm regime that enables in situ correction of the error in DNA extension measurement, and normalize

N-ethylmaleimide sensitive factor (NSF) is a key protein of intracellular membrane traffic. NSF is a highly conserved protein belonging to the ATPases associated with other activities (AAA+ proteins). AAA+ share common domains and all transduce ATP hydrolysis into major conformational movements that are used to carry out conformational work on client proteins. Together with its cofactor SNAP, NSF is specialized on disassembling highly stable SNARE complexes that form after each membrane fusion e

Receptor tyrosine kinase Mer (MerTK) has been shown to be highly expressed in Glioblastoma multiforme (GBM) in comparison to its healthy counterpart and is implicated in brain tumorigenesis. Clarifying the underlying mechanism of MerTK induced invasiveness would result in novel strategies to improve patient's response to chemotherapeutics. In vitro and in vivo assays were performed to examine the functional role of cancer stem sell (CSC) maintenance in MerTK associated invasiveness. In this arti

DNA barcoding provides a way to label a myriad of different biological molecules using the extreme programmability in DNA sequence synthesis. Fluorescence imaging is presumably the most easy-to-access method for DNA barcoding, yet large spectral overlaps between fluorescence dyes severely limit the numbers of barcodes that can be detected simultaneously. We here demonstrate the use of single-molecule fluorescence resonance energy transfer (FRET) to encode virtual signals in DNA barcodes using co

Heterogeneity in a tumor allows a small portion of cancer cells to survive and regrow upon targeted cancer therapy, eventually leading to cancer relapse. Such drug-resistant cells often exhibit dynamic adaptation of their signaling pathways at the level of protein-protein interactions (PPIs). To probe the rewiring of signaling pathways and the heterogeneity across individual cancer cells, we developed a single-cell version of the co-immunoprecipitation (co-IP) analysis that examines the amount a

Cancer cells actively release extracellular vesicles (EVs) as important carriers of cellular information to tumor microenvironments. Although the composition and quantity of the proteins contained in EVs are characterized, it remains unknown how these proteins in EVs are related to those in the original cells at the functional level. With epidermal growth factor receptor (EGFR) in lung adenocarcinoma cells as a model oncoprotein, it is studied how distinct types of EVs, microvesicles and exosome