Gyudo Lee

Mapping the surface charge distribution of amyloid fibril

It is of high importance to measure and map the surface charge distribution of amyloids, since electrostatic interaction between amyloidogenic proteins and biomolecules plays a vital role in amyloidogenesis. In this work, we have measured and mapped the surface charge distributions of amyloids (i.e., β-lactoglobulin fibril) using Kelvin probe force microscopy. It is shown that the surface charge distribution is highly dependent on the conformation of amyloids (e.g., the helical pitch of amyloid fibrils) as well as the pH of a solvent.

Single-Molecule Recognition of Biomolecular Interaction via Kelvin Probe Force Microscopy

We report the scanning probe microscope (SPM)-based single-molecule recognition of biomolecular interactions between protein kinase and small ligands (i.e., ATP and Imatinib). In general, it is difficult to sense and detect the small ligands bound to protein kinase (at single-molecule resolution) using a conventional atomic force microscope (AFM) due to the limited resolution of conventional AFM for detecting the miniscule changes in molecular size driven by ligand binding. In this study, we have demonstrated that Kelvin probe force microscopy (KPFM) is able to articulate the surface potential of biomolecules interacting with ligands (i.e., the protein kinase-ATP interactions and inhibition phenomena induced by antagonistic molecules) in a label-free manner. Furthermore, measured surface potentials for biomolecular interactions enable quantitative descriptions on the ability of protein kinase to interact with small ligands such as ATP or antagonistic molecules. Our study sheds light on KPFM that allows the precise recognition of single-molecule interactions, which opens a new avenue for the design and development of novel molecular therapeutics.

Real‐Time Quantitative Monitoring of Specific Peptide Cleavage by a Proteinase for Cancer Diagnosis

.…(MMPs;greenmissilesinthepicture)expressedonacancercellsurfacecanbesensedby a resonant cantilever device (satellite arm in the picture), as J. Yang, D. S. Yoon, T.Kwon et al. report in their Communication on page 5837 ff. Active MMPs attack thepeptide sequence that is immobilized on the cantilever surface. The peptide cleavageleads to an increase in the resonant frequency of the cantilever, owing to a decrease inthe mass of immobilized peptide.

Hyaluronic acid receptor-targetable imidazolized nanovectors for induction of gastric cancer cell death by RNA interference

We have developed a nanovector consisting of hyaluronic acid (HA) and poly-L-lysine-graft-imidazole (PLI)-based polyplexes containing Bcl-xL-specific shRNA-encoding plasmid DNA (HA/PLI/pDNA) for CD44 targeted gastric cancer therapy. The prepared ternary polyplexes have a negative surface charge of -24 mV and a size of approximately 100 nm at an N/P ratio of 5 with HA/PLI molar ratio of 0.03. Gel electrophoresis and cell viability experiments demonstrated that the ternary polyplexes showed high stability and no cytotoxicity due to the anchored HA molecules on the surface of PLI/pDNA binary polyplexes. Selective cancer cell death was achieved by CD44-mediated gene delivery and the internalized gene was effectively escaped from endosomes due to the buffering capacity of imidazole groups in an acidic environment. These nanovectors may be highly efficient gene delivery tools that allow the selective destruction of metastatic gastric cancer cells.

Aptamer-functionalized nano-pattern based on carbon nanotube for sensitive, selective protein detection

We have developed a horizontally aligned carbon nanotube sensor that enables not only the specific detection of biomolecules with ultra-sensitivity, but also the quantitative characterization of binding affinity between biomolecules and/or interaction between a carbon nanotube and a biomolecule, for future applications in early diagnostics. In particular, we have fabricated horizontally aligned carbon nanotubes, which were functionalized with specific aptamers that are able to specifically bind to biomolecules (i.e. thrombin). Our detection system is based on scanning probe microscopy (SPM) imaging for horizontally aligned aptamer-conjugated carbon nanotubes (ACNTs) that specifically react with target biomolecules at an ultra-low concentration. It is shown that the binding affinity between thrombin molecule and ACNT can be quantitatively characterized using SPM imaging. It is also found that the smart carbon nanotube sensor coupled with SPM imaging permits us to achieve the high detection sensitivity even up to ∼1 pM, which is much higher than that of other bioassay methods. Moreover, we have shown that our method enables a quantitative study on small molecule-mediated inhibition of specific biomolecular interactions. In addition, we have shown that our ACNT-based system allows for the quantitative study of the effect of chemical environment (e.g. pH and ion concentration) on the binding affinity. Our study sheds light on carbon nanotube sensor coupled with SPM imaging, which opens a new avenue to early diagnostics and drug screening with high sensitivity.

Nanomechanical characterization of chemical interaction between gold nanoparticles and chemical functional groups

We report on how to quantify the binding affinity between a nanoparticle and chemical functional group using various experimental methods such as cantilever assay, PeakForce quantitative nanomechanical property mapping, and lateral force microscopy. For the immobilization of Au nanoparticles (AuNPs) onto a microscale silicon substrate, we have considered two different chemical functional molecules of amine and catecholamine (here, dopamine was used). It is found that catecholamine-modified surface is more effective for the functionalization of AuNPs onto the surface than the amine-modified surface, which has been shown from our various experiments. The dimensionless parameter (i.e., ratio of binding affinity) introduced in this work from such experiments is useful in quantitatively depicting such binding affinity, indicating that the binding affinity and stability between AuNPs and catecholamine is approximately 1.5 times stronger than that between amine and AuNPs. Our study sheds light on the experiment-based quantitative characterization of the binding affinity between nanomaterial and chemical groups, which will eventually provide an insight into how to effectively design the functional material using chemical groups.

Characterization of the regrowth behavior of amyloid-like fragmented fibrils decomposed by ultrasonic treatment

Ultrasonic treatment (UST) has been used not only to accelerate protein fibril growth and amplify infectious prion proteins from biological fluids, but also to break amyloid-like fibrils for treatment. Despite the applicability of UST to clinical treatment, both the decomposition characteristics of fibrils and the regrowth mechanisms of the decomposed fibrils remain unclear. Here, we report UST-driven decomposition of amyloid-like fibrils into shorter fibrils and the principles of fibril regrowth via conventional heating. Short fibrils decomposed by UST can be reconstructed into mature fibrils with the addition of monomeric protein, but the regrowth of short fibrils rarely occurs in the absence of monomeric protein. Interestingly, the reconstructed fibrils possess electric properties similar to those of the original fibrils. We propose a model that describes the regrowth process of amyloid-like fibrils decomposed by UST, which sheds light on future biomedical applications of UST for the treatment of amyloidogenic diseases.

Carbon Nanotube-Patterned Surface-Based Recognition of Carcinoembryonic Antigens in Tumor Cells for Cancer Diagnosis

It has been of high significance to devise a biochemical analytical tool kit enabling the detection of few circulating tumor cells (CTCs) for early diagnosis of cancer. Despite recent effort made to detect few CTCs, it is still challenging to sense such cells with their low concentration and/or the minute amount of marker proteins expressed on few CTCs. In this work, we report the label-free recognition of carcinoembryonic antigens (CEAs) expressed on few CTCs by using a carbon nanotube (CNT) sensor coupled with scanning probe microscopy imaging for cancer diagnosis. It is shown that a CNT-patterned surface is able to specifically capture the CEA molecules in the whole cell lysate of CTCs with their concentration even up to 10(-3) cells/mL. Our work sheds light on our bioassay based on a CNT-patterned surface for highly sensitive, label-free detection of marker proteins expressed on few tumor cells, which may open a new avenue in early diagnosis of cancer by providing a novel biochemical analysis tool kit.

Single-step electropolymerization patterning of a polypyrrole nanowire by ultra-short pulses via an AFM cantilever

Conducting polymers (CPs) have attracted a great deal of attention due to their unique properties; these properties are useful in implementing various functional devices, such as memory, and chemical and biological sensors. In particular, the nanopatterning of CPs is a key technology that will accelerate the adoption of CPs in fabricating nanoscaled multifunctional devices. This paper presents an innovative technique for forming polypyrrole nanowire (PPy-NW) patterns, without any additional pretreatment on the gold surface, using atomic force microscopy (AFM) and ultra-short pulse voltage. Applying the ultra-short pulse voltage to the AFM tip has the following advantage: since the electrochemical current is extremely localized around the tip, the successful formation of CP nanowires results. This is because the pulse width is much shorter than the resistor-capacitor (RC) time constant of the equivalent electrochemical circuit of our experimental set-up. This paper provides systematic results regarding the dimensional variation of the PPy-NW patterns produced by varying the electrical conditions of the ultra-short pulse, such as the pulse amplitude, width, and frequency. The results show that use of an ultra-short pulse is essential in fabricating PPy-NW patterns. Additionally, an ultra-short pulse offers excellent pattern controllability for both width (353 nm ∼ 3.37 µm) and height (2.0 ∼ 88.3 nm).

Experimental and numerical study of electrochemical nanomachining using an AFM cantilever tip

We fabricated nanopatterns on Cu thin films via an electrochemical route using an atomic force microscope (AFM). Experimental results were compared with an equivalent electrochemical circuit model representing an electrochemical nanomachining (ECN) technique. In order to precisely construct the nanopatterns, an ultra-short pulse was applied onto the Cu film through the AFM cantilever tip. The line width of the nanopatterns (the lateral dimension) increased with increased pulse amplitude, on-time, and frequency. The tip velocity effect on the nanopattern line width was also investigated. The study described here provides important insight for fabricating nanopatterns precisely using electrochemical methods with an AFM cantilever tip.