發展奈米探針用以快速和專一性純化硫基亞硝基化胜肽

楊牧軒

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發展奈米探針用以快速和專一性純化硫基亞硝基化胜肽

Development of Affinity Nano Probes for Rapid and Specific Enrichment of S-nitrosylated Peptides

楊牧軒 , Masters　　Advisor：林俊成　　

繁體中文

硫基亞硝基化胜肽；奈米探針；親合性探針；磁性奈米粒子； S-nitrosylated Peptides ； Nano Probes ； Affinity probes ； magnetic nanoparticle

Abstract │ Reference (78) │ Altmetrics

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Nitric oxide (NO) is covalently attached to cysteine thiols of proteins resulting in the formation of S-nitrosothiols (RSNOs), and regulation of protein activity and function in a wide range of physiological processes. The objectives of this thesis are development of probes for the enrichment of S-nitrosylated peptides. We conjugated two types of thioester phosphine ligand to either magnetic nanoparticle or agarose bead, respectively for identification and purification of RSNOs. Furthermore,  through traceless reductive ligation of S-nitrosothiol mechanism the unstable primary RSNOs were converted to stable disulfide-iminophosphorane products. The S-nitrosylated PTP1B peptide mixed with tryptic BSA mixture was successfully captured by developed  probe and identified by MALDI-TOF. In addition, the agarose beads show less non-specific interaction to tryptic BSA peptide. The enrichment of S-nitrosylated peptides from COS-7 cell lysate was achieved by 2nd_TEP@agarose. Thus, This strategy is a potential tool for investigate of S-nitrosylation proteins. Furthermore, we synthesized other type of thioester phosphine ligand by changing the original structure and investigate the effect of probe structure on the reactivity to RSNO.

Reference ( 78 ) 〈TOP〉

1. Arnold, W. P.; Mittal, C. K.; Katsuki, S.; Murad, F., Nitric oxide activates guanylate cyclase and increases guanosine 3':5'-cyclic monophosphate levels in various tissue preparations. Proc. Natl. Acad. Sci. U. S. A. 1977, 74, 3203-3207.
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2. Bin-Nun, A.; Schreiber, M. D., Role of iNO in the modulation of pulmonary vascular resistance. J. Perinatol. 2008, 28, 84-92.
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3. Furchgott, R. F.; Zawadzki, J. V., The obligatory role of endothelial cells in the relaxation of arterial smooth muscle by acetylcholine. Nature 1980, 288 , 373-376.
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4. Ignarro, L. J.; Buga, G. M.; Wood, K. S.; Byrns, R. E.; Chaudhuri, G., Endothelium-derived relaxing factor produced and released from artery and vein is nitric oxide. Proc. Natl. Acad. Sci. U. S. A. 1987, 84, 9265-9269.
連結：
5. Odell, T. J.; Hawkins, R. D.; Kandel, E. R.; Arancio, O., Tests of the Roles of 2 Diffusible Substances in Long-Term Potentiation - Evidence for Nitric-Oxide as a Possible Early Retrograde Messenger. Proc. Natl. Acad. Sci. U. S. A. 1991, 88, 11285-11289.
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