J.O. Lee, G.B. Kim, Y.-P. Kim
Hanyang University, Korea
pp. 133 - 136
Keywords: gold nanoparticle, protein phosphatase, zinc, histidine tag
Since gold nanoparticle (AuNP)-based colorimetric assays have been preferred for enzyme activity assay, a few of attempts have been designed for the simple and rapid detection of PP activity. However, they still have drawbacks including antibody-dependent variations and complicate synthesis of rare chemicals to specifically capture phosphorylated amino acids. To tackle this problem, here we design a simple AuNP colorimetric assay of PP activity using His6-tagged peptides and Zn(II) ions, which does not require antibody and rare chemical. Among metal ions including nickel (II), copper(II), cobalt(II), magnesium (II), manganese (II), and zinc(II), only zinc (II) triggered a strong association between His6-tagged phosphopeptides and carboxyl groups on the surface of AuNPs (20 nm in core diameter), leading to the self-assembly of AuNPs and subsequently their color changes. As a result, the PP activity was easily quantified by the extinction ratio (E520/E700) of the colloidal gold assembly, where there was a linear correlation between the PP concentration and the extinction ratio. Most importantly, this method was successfully used for analysing protein phosphatase 2A (PP2A) activity in immunoprecipitated plant extracts. We anticipate that out method will find the applications to monitor the activities of various PPs and their inhibition in a rapid format.