Bergenshalvøens Kommunale Kraftselskap (Norway)

A simple, colorimetric method is developed for the determination of deoxyribonuclease I (DNase I) activity based on the novel finding that DNase I can promote the photoinduced synthesis of gold nanoparticles (AuNPs). In the absence of DNase I, a phosphorothioate (PS) DNA probe remains intact and captures Au(iii) through a strong Au-thiol interaction, which prevents the photoinduced synthesis of AuNPs, leaving the sample in a colorless state. On the other hand, in the presence of DNase I, the PS DNA probe is cleaved into small fragments that are removed via a simple purification process. The resulting solution, after the incubation with HAuCl4 and threonine (Thr), forms AuNPs by UV light irradiation with the aid of Thr which acts as a catalyst for the Au(iii) reduction process. As a result, a red-colored suspension is produced. By monitoring the color changes of the samples with the naked eye, the DNase I activity was conveniently determined. In addition, the clinical utility of this simple, yet highly efficient colorimetric strategy was verified by reliably quantifying the DNase I activities in a bovine urine sample. Importantly, the working principle designed for the determination of DNase I activity was successfully expanded for the detection of target nucleic acids, ensuring the universal applicability of the developed assay system.

We herein describe a novel impedimetric method to determine alkaline phosphatase (ALP) activity based on the Cu2+-mediated oxidation of ascorbic acid on a specific DNA probe-modified electrode. In this method, pyrophosphate (PPi) capable of complexing with Cu2+ is employed as a substrate of the ALP enzyme. In the presence of ALP, PPi is hydrolyzed to phosphate (Pi), which is not able to entrap Cu2+. The free Cu2+ are specifically bound to a poly-thymine DNA probe immobilized on the electrode surface and reduced to form copper nanoparticles by a concomitant oxidation of ascorbic acid. As a result, the oxidation products of ascorbic acid are accumulated on the electrode surface, which consequently increase electron transfer resistance (Ret) by interrupting the electron transfer on the electrode. On the other hand, in the absence of ALP, PPi remains intact to effectively capture Cu2+, consequently preventing the oxidation of ascorbic acid and the subsequent increase of Ret. Based on this design principle, the change in Ret, which is proportional to ALP activity, was measured by electrochemical impedance spectroscopy (EIS) and ALP activities were successfully determined down to 6.5 pM (7.2 U L−1) with excellent selectivity.

A rapid and label-free strategy for sensitive detection of Hg2+ is developed based on target-triggered exponential strand displacement amplification (eSDA).

Human SIRT1 controls various physiological responses including cell fate, stress, and aging, through deacetylation of its specific substrate protein. In processing DNA damage signaling, SIRT1 attenuates a cellular apoptotic response by deacetylation of p53 tumor suppressor. The present study shows that, upon exposure to radiation, SIRT1 could enhance DNA repair capacity and deacetylation of repair protein Ku70. Ectopically over-expressed SIRT1 resulted in the increase of repair of DNA strand breakages produced by radiation. On the other hand, repression of endogenous SIRT1 expression by SIRT1 siRNA led to the decrease of this repair activity, indicating that SIRT1 can regulate DNA repair capacity of cells with DNA strand breaks. In addition, we found that SIRT1 physically complexed with repair protein Ku70, leading to subsequent deacetylation. The dominant-negative SIRT1, a catalytically inactive form, did not induce deacetylation of Ku70 protein as well as increase of DNA repair capacity. These observations suggest that SIRT1 modulates DNA repair activity, which could be regulated by the acetylation status of repair protein Ku70 following DNA damage.