量子ドット技術を用いた複数のタンパク質翻訳後修飾の同時検出

抄録

The number of protein isoforms in the human proteome has known to be much higher than the number of genes in the human genome. This is in large part due to posttranslational modifications (PTMs) of proteins. Knowledge of these PTMs is extremely important because they may altar physical and chemical properties, folding, complex formation, stability, activity, and consequently, function of the proteins. The Quantum dots are high-intensity photostable fluorophores that have several distinct advantages over standard organic fluorescent dyes. These nanocrystals (commercially available as Qdots from Invitrogen; Carlsbad, CA, USA) consist of a semiconductor core of cadmium selenide (CdSe) coated with a shell of zinc sulfide(ZnS). An additional polymer layer over the Qdot enhances water solubility and enables conjugation to streptavidin or other biomolecules such as IgG. Slight changes in the size of the Qdot semiconductor core change the emission spectra, resulting in a redder color as the size of the core increases. The ability to synthesize different populations of quantum dots with narrow, symmetric and strong emission spectra permits multiplexing, a property that is important for simultaneous detection of several types of PTMs. With the Qdot technology, we developed a new proteomic method for simultaneous detection of various PTMs of proteins on a single Western or immunoblotting analysis. We applied this method to examine the changes of the PTMs of proteins from various tissues and cells in a diabetes model Otsuka Long-Evans Tokushima Fatty (OLETF) rat. Protein carbonyls, ubiquitination and advanced glycation end products (AGEs), were detected simultaneously on a single blot. The advantages of high sensitivity, multiplex labeling, photo- and chemical stability of the Qdots will accelerate information flow in proteomics.