Browsing by Author "Wang, Eng-Chi"

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Alignment of Stretchable Nanoparticle Chains with Tunable Optical Properties Formed from Molecular Machinery
(2013-01-01)
Lin, Ming-Yu
;
Lu, Yen-Pei
;
Yang, Yuh-Shyong
;
Chen, Hsuen-Li
;
Yang, Chih-Hui
Control orientation and interparticle spacing of self-assembled nanoparticles can provide versatile functional materials for optical and electronic applications. In this study, we have obtained a glass substratum with tunable optical properties using flowing control and bimolecular competition. The surface of glass was modified from immobilized stretchable gold nanoparticle chains. The long and periodic single-stranded DNA nanotemplates were prepared using Phi29 DNA polymerase molecular machinery according to precise molecular biology regulation. Gold nanoparticle (Au NP) docked at the default sites of the nanotemplates and optical anisotropic absorption were enhanced by approximately 55 % by using tunable, flexible, and oriented one-dimensional metallic nanostructures.
Azide-alkyne cycloaddition for universal post-synthetic modifications of nucleic acids and effective synthesis of bioactive nucleic acid conjugates
(2014)
Su, Yu-Chih
;
Lo, Yu-Lun
;
Hwang, Chi-Ching
;
Wang, Li-Fang
;
Wu, Min Hui
The regioselective post-synthetic modifications of nucleic acids are essential to studies of these molecules for science and applications. Here we report a facile universal approach by harnessing versatile phosphoramidation reactions to regioselectively incorporate alkynyl/azido groups into post-synthetic nucleic acids primed with phosphate at the 5\' termini. With and without the presence of copper, the modified nucleic acids were subjected to azide-alkyne cycloaddition to afford various nucleic acid conjugates including a peptide-oligonucleotide conjugate (POC) with high yield. The POC was inoculated with human A549 cells and demonstrated excellent cell-penetrating ability despite cell deformation caused by a small amount of residual copper chelated to the POC. The combination of phosphoramidation and azide-alkyne cycloaddition reactions thus provides a universal regioselective strategy to post-synthetically modify nucleic acids. This study also explicated the toxicity of residual copper in synthesized bioconjugates destined for biological systems.
Effective and site-specific phosphoramidation reaction for universally labeling nucleic acids
(2014-03-15)
Su, Yu-Chih
;
Chen, Hsing-Yin
;
Ko, Ni Chien
;
Hwang, Chi-Ching
;
Wu, Min Hui
Here we report efficient and selective postsynthesis labeling strategies, based on an advanced phosphoramidation reaction, for nucleic acids of either synthetic or enzyme-catalyzed origin. The reactions provided phosphorimidazolide intermediates of DNA or RNA which, whether reacted in one pot (one-step) or purified (two-step), were directly or indirectly phosphoramidated with label molecules. The acquired fluorophore-labeled nucleic acids, prepared from the phosphoramidation reactions, demonstrated labeling efficacy by their F/N ratio values (number of fluorophores per molecule of nucleic acid) of 0.02-1.2 which are comparable or better than conventional postsynthesis fluorescent labeling methods for DNA and RNA. Yet, PCR and UV melting studies of the one-step phosphoramidation-prepared FITC-labeled DNA indicated that the reaction might facilitate nonspecific hybridization in nucleic acids. Intrinsic hybridization specificity of nucleic acids was, however, conserved in the two-step phosphoramidation reaction. The reaction of site-specific labeling nucleic acids at the 5'-end was supported by fluorescence quenching and UV melting studies of fluorophore-labeled DNA. The two-step phosphoramidation-based, effective, and site-specific labeling method has the potential to expedite critical research including visualization, quantification, structural determination, localization, and distribution of nucleic acids in vivo and in vitro. (C) 2013 Elsevier Inc. All rights reserved.
In Vitro Selection and Characterization of a Novel Zn(II)-Dependent Phosphorothiolate Thiolesterase Ribozyme
(2012-01-10)
Wang, Tzu-Pin
;
Su, Yu-Chih
;
Chen, Yi
;
Liou, Yi-Ming
;
Lin, Kun-Liang
Here we present the in vitro selection of a novel ribozyme specific for Zn(2+)-dependent catalysis on hydrolysis of a phosphorothiolate thiolester bond. The ribozyme, called the TW17 ribozyme, was evolved and selected from an artificial RNA pool covalently linked to a biotin-containing substrate through the phosphorothiolate thiolester bond. The secondary structure for the evolved ribozyme consisted of three major helices and three loops. Biochemical and chemical studies of ribozyme-catalyzed reaction products provided evidence that the ribozyme specifically catalyzes hydrolysis of the phosphorothiolate thiolester linkage. A successful ribozyme construct with active catalysis in trans further supported the determined ribozyme structure and indicated the potential of the ribozyme for multiple-substrate turnover. The ribozyme also requires Zn(2+) and Mg(2+) for maximal catalysis. The TW17 ribozyme, in the presence of Zn(2+) and Mg(2+), conferred a rate enhancement of at least S orders of magnitude when compared to the estimated rate of the uncatalyzed reaction. The ribozyme completely lost catalytic activity in the absence of Zn(2+), like Zn(2+)-dependent protein hydrolases. The discovery and characterization of the TW17 ribozyme suggest additional roles for Zn(2+) in ribozyme catalysts.
Successful preparation and characterization of biotechnological grade agarose from indigenous Gelidium amansii of Taiwan
(2012-03-01)
Wang, Tzu-Pin
;
Chang, Li-Lin
;
Chang, Sheng-Nan
;
Wang, Eng-Chi
;
Hwang, Long-Chih
This paper reports the first successful preparation of biotechnological grade agarose from Gelidium amansii found in Taiwan. The scale-efficiency preparation was achieved by shortening EDTA treatment time through dispersing G. amansii agar in water in the presence of heat and EDTA, removing agaropectin impurity with a heat-compatible and strong-anion exchange resin, and precipitating agarose with a cost-effective isopropanol method. The yield of agarose from prepared G. amansii agar was 11.3%. The acquired agarose has a gel strength of 853 g cm(-2), a sulfate content of 0.14%, a pyruvate content of 1.03%, a degree of electroendosmosis of 0.16 and very limited binding affinity to DNA. The excellent properties of agarose from G. amansii of Taiwan confirm its potential diverse biotechnological applications. This innovative agarose preparation method with the significantly improved scale-efficiency can be modified for large-scale preparation of agarose for use in biotechnological industry and biochemical research. (C) 2011 Elsevier Ltd. All rights reserved.