A glycine spacer improved peptidyl-nuclear-localized efficiency for fluorescent imaging nuclear H2O2

Abstract

H2O2 generated aberrantly can attack DNA in nuclei and result in oxidative DNA damage, which has been associated with many serious diseases. Therefore, it is crucial to monitor nuclear H2O2 levels. However, the vast majority of fluorescence sensors can not localize in the nucleus. In the present study, we empolyed glycine (the simplest amino acids) as a spacer to lengthen distance between N50 and NP1. Cu2+-free click reaction (through inserting dibenzocyclooctyne-maleimide) was used to connect the two parts in situ. Three spacer sequences were tested: C, CGGGGG, and CGGGGGGGGGG, and three new NLS peptides (connection spacer sequence to N50) were generated: pep5, pep6 and pep7. The fluorescence imaging results showed the nucleus-targeting efficancy of the three peptides, especially pep6, were imporved. The calculated binding energies by molecular docking simulations with importin α5 (a member of importin) were pep6 < pep5 < pep7, consistent with the fluorescence imaging results. Importantly, the probe system consisting of pep6 and NP1 was used for imaging the increase of nuclear H2O2 level upon stimulation.