surface-passivation-for-single-molecule-protein-studies

Surface Passivation for Single-molecule Protein Studies

Single-molecule fluorescence spectroscopy has proven to be instrumental in understanding a wide range of biological phenomena at the nanoscale. Important ...

Seoul National University

A full-length cDNA of the Type I procollagen alpha1 [pro-alpha1(I)] chain (4388 bp), coding for 1463 amino acid residues in the total length, was determined by RACE PCR using a cDNA library constructed from 4-week embryo of the skate Raja kenojei. The helical region of the skate pro-alpha1(I) chain consisted of 1014 amino acid residues - the same as other fibrillar collagen alpha chains from higher vertebrates. Comparison on denaturation temperatures of Type I collagens from the skate, rainbow trout (Oncorhynchus mykiss) and rat (Rattus norvegicus) revealed that the number of Gly-Pro-Pro and Gly-Gly in the alpha1(I) chains could be directly related to the thermal stability of the helix. The expression property of the skate pro-alpha1(I) chain mRNA and phylogenetic analysis with other vertebrate pro-alpha1(I) chains suggested that skate pro-alpha1(I) chain could be a precursor form of the skate Type I collagen alpha1 chain. The present study is the first evidence for the primary structure of full-length pro-alpha1(I) chain in an elasmobranch.

cDNA cloning and characterization of Type I procollagen alpha1 chain in the skate Raja kenojei.

The organic solute and steroid transporter (OST/Ost) is a unique membrane transport protein heterodimer composed of subunits designated alpha and beta, that transports conjugated steroids and prostaglandin E(2) across the plasma membrane. Ost was first identified in the liver of the cartilaginous fish Leucoraja erinacea, the little skate, and subsequently was found in many other species, including humans and rodents. The present study describes the isolation of a new cell line, LEE-1, derived from an early embryo of L. erinacea, and characterizes the expression of Ost in these cells. The mRNA size and amino acid sequence of Ost-beta in LEE-1 were identical to that previously reported for Ost-beta from skate liver, and the primary structure was identical to that of the spiny dogfish shark (Squalus acanthias) with the exception of a single amino acid. Ost-beta was found both on the plasma membrane and intracellularly in LEE-1 cells, consistent with its localization in other cell types. Interestingly, arachidonic acid, the precursor to eicosanoids, strongly induced Ost-beta expression in LEE-1 cells and a lipid mixture containing arachidonic acid also induced Ost-alpha. Overall, the present study describes the isolation of a novel marine cell line, and shows that this cell line expresses relatively high levels of Ost when cultured in the presence of arachidonic acid. Although the function of this transport protein in embryo-derived cells is unknown, it may play a role in the disposition of eicosanoids or steroid-derived molecules.

Arachidonic acid-induced expression of the organic solute and steroid transporter-beta (Ost-beta) in a cartilaginous fish cell line.

Stem cells from cartilaginous and bony fish.

Natural products in Chonnam, Korea were screened via anti-angiogenesis experiments, and 1 candidate product was identified, Corni fructus, which exerted dose-dependent inhibitory effects against angiogenesis, adipogenesis, and cell adhesion. C. fructus extract (CFE) exhibits an angiogenesis inhibitory effect superior to that of the EGCG from green tea leaves. The expression level of angiogenesis and adipogenesis-related signal molecules in the western blotting was reduced by increasing the amount of added CFE. Moreover, a diet supplemented with CFE was deemed more effective in inducing weight loss in LB mice than a representative synthetic diet drug, orlistat, which incidently caused the side effect of denuding the mice of their hair. These results indicate that C. fructus may prove to be a useful anti-adipogenic compound, and these in vitro results may be reflected later under in vivo conditions.

Inhibitory effects of corni fructus extract on angiogenesis and adipogenesis.

An ideal surface-enhanced Raman scattering (SERS) nanostructure for sensing and imaging applications should induce a high signal enhancement, generate a reproducible and uniform response, and should be easy to synthesize. Many SERS-active nanostructures have been investigated, but they suffer from poor reproducibility of the SERS-active sites, and the wide distribution of their enhancement factor values results in an unquantifiable SERS signal. Here, we show that DNA on gold nanoparticles facilitates the formation of well-defined gold nanobridged nanogap particles (Au-NNP) that generate a highly stable and reproducible SERS signal. The uniform and hollow gap (∼1 nm) between the gold core and gold shell can be precisely loaded with a quantifiable amount of Raman dyes. SERS signals generated by Au-NNPs showed a linear dependence on probe concentration (R(2) > 0.98) and were sensitive down to 10 fM concentrations. Single-particle nano-Raman mapping analysis revealed that >90% of Au-NNPs had enhancement factors greater than 1.0 × 10(8), which is sufficient for single-molecule detection, and the values were narrowly distributed between 1.0 × 10(8) and 5.0 × 10(9).

Highly uniform and reproducible surface-enhanced Raman scattering from DNA-tailorable nanoparticles with 1-nm interior gap.

Tooth loss evokes functional and esthetic problems, and adjacent teeth can move if they are not corrected in a timely manner, rendering the problems of restoring the dentition more complex. If the opposing tooth is consequently extruded, several treatment modalities may be adopted, such as occlusal adjustment, crown prosthesis, orthodontic treatment, and segmental osteotomy. Segmental osteotomy is the most aggressive and invasive treatment for restoring a problematic occlusal plane due to the complications, such as hemorrhage, loss of tooth vitality, or necrosis of the segment. Piezoelectric devices have recently been used in oral surgery and implant dentistry because they preserve the soft tissue. In this report, we describe a bilateral posterior segmental osteotomy procedure to correct a collapsed occlusal plane with sinus mucosa elevation, using piezoelectric devices to preserve the integrity of the sinus membrane and the posterior superior alveolar artery.

Posterior maxillary segmental osteotomy concomitant with sinus lift using a piezoelectric device.

The plasmonic properties of metallic nanoparticles (NPs) such as Au and Ag NPs and the plasmonic coupling between them are of enormous interest for their strong and controllable optical signal enhancement and manipulation capabilities. The strong optical properties of these plasmonic structures are promising for various biosensing applications, but the widespread use of these structures is limited largely due to the absence of high-yield synthetic method for targeted nanoprobes with nanometer precision and the poor understanding of the plasmonics of these structures. DNA is a promising material that can be used as both specific biorecognition and versatile synthetic template in forming and controlling plasmonic nanostructures and their aggregations. In this article, we provide an overview and perspective of recent advances in the use of DNA-tailored plasmonic nanostructures in biosensing applications.

DNA-tailored plasmonic nanoparticles for biosensing applications.

Here, we highlight the strategies for the synthesis and tuning of a variety of metal nanostructures and nanoassemblies with oligonucleotides and their applications. We have discussed the importance of and need for various metal nanostructures and the role and suitability of DNA in building these nanostructures and nanoassemblies. A large part of this article is devoted to the discussion of DNA-mediated synthetic methods for metal nanostructures. The synthetic strategies are categorized into two groups - strategy 1 that uses DNA as the ligand for metal nanoparticles and subsequent assembly or modification and strategy 2 that uses DNA template-directed assemblies of metal nanoparticles.

Jae-Ho Hwang

Department of Chemistry

Seoul National University

Surface Passivation for Single-molecule Protein Studies