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Research Overview
The Lin research group studies the dynamic behavior of biomolecules—such as proteins, nucleic acids, and biological membranes—at the single-molecule level. Biological function is inherently dynamic, involving conformational changes, diffusion, self-assembly, and molecular interactions. To capture these events in action, we develop and apply high-speed atomic force microscopy (HS-AFM) and integrate our results with complementary techniques, including fluorescence microscopy, electron microscopy, and electrophysiology.
We aim to visualize structural and functional dynamics in physiologically-relevant environments with nanometer spatial and millisecond temporal resolution. Our multidisciplinary approach leverages expertise in:
HS-AFM imaging and force spectroscopy
Single-molecule biophysics and structural biology
Image processing, machine learning, and kinetic modeling
Novel instrument and method development
By pushing the frontiers of real-time molecular imaging, we aim to uncover the mechanisms that drive complex biological processes.
High-Speed Atomic Force Microscopy
HS-AFM has revolutionized single-molecule biophysics and structural biology by providing direct insights into protein dynamics in native-like environments, membrane protein function & lipid interactions, protein & nucleic acid interactions, and mechanotransduction & biomolecular mechanics. Unlike conventional AFM, which lacks the speed to capture rapid molecular events, HS-AFM uniquely reveals dynamic biological phenomena in real time and under near-physiological conditions.
This transformative technology allows us to observe:
Transient molecular interactions
Protein conformational changes
Assembly and disassembly of biomolecular complexes
Structural transitions of biomolecules at different functional states
These approaches enable us to uncover the physical principles and mechanisms driving complex biological processes, advancing both fundamental science and biomedical research.
HS-AFM operates by scanning a sharp nanoscale probe across a sample surface, detecting changes in height and producing high-resolution topographical images.
How HS-AFM Compares to Other Structural and Functional Characterization Tools?
High-Speed Atomic Force Microscopy (HS-AFM) occupies a unique niche in structural and functional characterization, complementing other high-resolution techniques like cryo-electron microscopy (cryo-EM), X-ray crystallography, and fluorescence microscopy. While cryo-EM and X-ray crystallography excel at providing atomic-level static structures, HS-AFM uniquely captures dynamic molecular processes in real time and under near-physiological conditions. Unlike fluorescence microscopy, which often relies on labels or dyes, HS-AFM provides label-free imaging, preserving the native state of biomolecules.
Moreover, HS-AFM stands out for its ability to probe mechanical properties and molecular interactions at the single-molecule level, offering insights into force-induced conformational changes and protein-lipid dynamics. Its compatibility with aqueous environments allows for direct observation of biomolecular assemblies, bridging the gap between static structural data and dynamic functional studies. By integrating HS-AFM with computational modeling and complementary techniques, our research group leverages the strengths of these tools to uncover the intricate mechanisms driving molecular function and assembly.
Protein Dynamics in Physiological Conditions
Our group applies HS-AFM to explore the dynamic behavior of proteins in their native-like environments. By capturing real-time conformational changes and transient states, we aim to elucidate the molecular mechanisms that govern essential protein functions, including folding, ligand binding, and enzymatic activity. Our research focuses on understanding how proteins respond to physiological stimuli and interact with other biomolecules in complex environments. Through advanced HS-AFM imaging techniques and methodological innovations, we bridge the gap between structural and functional studies, providing new insights into protein dynamics and their roles in cellular processes.
Realtime dynamics of Annexin-V lattice on membrane with newly developed image reconstruction technique.
Membrane Proteins and Their Interactions with Lipids
Membrane proteins (transmembrane and membrane-binding proteins ) play an essential role in maintaining the homeostasis of cells by functioning as transporters for signal transaction and energy conversion, among other functions. Therefore, knowledge of the atomic-resolution structures of membrane proteins is crucial to understanding their functions.
Our research investigates the structure and dynamics of membrane proteins and their interactions with surrounding lipid bilayers. Using HS-AFM, we directly visualize real-time conformational changes, lipid-protein interactions, and membrane remodeling events at nanometer resolution. Our research aims to uncover how the lipid bilayer influences protein function and how proteins, in turn, modulate membrane properties, providing key insights into their roles in cellular physiology and disease mechanisms.
Membrane-reconsituted OmpF trimers viewed from extracellular side (Left) and periplasmic side (Right).
Protein-Nucleic Acid Interactions
Our research leverages HS-AFM to study the dynamic interactions between proteins and nucleic acids, including DNA and RNA. Protein-nucleic acid complexes play a central role in essential cellular processes, including replication, transcription, and repair. We visualize these processes in real time, capturing transient conformational states and tracking the movement of proteins along nucleic acid strands. This allows us to investigate mechanisms such as DNA unwinding by helicases, polymerase activity, and nucleosome remodeling at nanometer resolution. By combining HS-AFM with complementary techniques, we aim to elucidate how proteins recognize, bind to, and modify nucleic acids, thereby shedding light on the molecular basis of gene regulation and genome stability.
DNA topological dynamics.
Instrument & Software Development
HS-AFM is a relatively young technique compared to established methods like electron and fluorescence microscopy, but it offers the unique ability to visualize biomolecules in real-time at the single-molecule level. To improve its speed, stability, resolution, and data analysis capabilities, we design advanced hardware, including next-generation feedback control systems and enhanced mechanical components, and develop algorithms—encompassing machine learning and AI tools—to automatically process and interpret HS-AFM data. Our research encompasses feedback control systems, image processing and data analysis, molecular recognition, and kinetic and biophysical modeling, to develop an integrated HS-AFM platform for rapid, accurate, and automated molecular studies.
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