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Inductively coupled plasma mass spectrometry (ICP-MS) is a powerful method for the matrix-independent quantitative analysis of target elements. Developed for the use in inorganic trace analysis, ICP-MS is nowadays emerging as a valuable tool for bioanalytical questions. Especially the use of ICP-MS for quantitative proteomics by measuring heteroatoms has gained popularity in the last decade, considering that established quantification methods like organic mass spectrometry depend on the existence of matched protein and peptide standards or labelling of the target protein. The need for reliable quantification of proteins is constantly growing, but only a limited number of well characterized and quantified protein standards are available so far. Not only in basic research, but also in a clinical context, accurately quantified, traceable protein standards are needed to ensure comparability of measurements between laboratories. One disease with a major impact on our ageing society is Alzheimer’s disease (AD), which is still challenging to diagnose. As this is also due to a lack in comparability and accuracy of existing biomarker assays, the community would greatly benefit from well quantified protein biomarker standards.
In this work, we applied isotope dilution analysis (IDA) using ICP-MS to quantify proteins of known stoichiometry via their sulfur content. Sulfur is present in two amino acids, cysteine and methionine, and hence exists in nearly all proteins. Simple strategies were employed for the detection of low molecular sulfur species to correct for sulfur contaminants and allow for reliable quantification of various proteins. We report the protein mass fractions with expanded uncertainties of a standard reference material and commercially available proteins determined by sulfur IDA. The herein developed method can be applied for the reliable and traceable quantification of pure proteins and will be used for the quantification of an AD biomarker. Our target is the tau protein, as brain load and distribution of tau is highly correlated with the clinical progression of AD.
The formation of DNA-protein complexes ocurrs during replication and repair within cells. They are assumed to modify the damage caused by ionization radiation during radaition therapy. Hereby the assumption is, that the underlying damaging channels in DNA and proteins are modified, especially when compared to single molecules.
Biopolymers are the building blocks of life. Their properties are exploited for material functionalization on the nanoscale in a flexible manner. An overview over current research activities in the field of sensing, nanostrcuturing, radiation damage measurements on DNA and proteins and microfluidics is given.
X-ray photoelectron-spectroscopy (XPS) allows simultaneous irradiation and damage monitoring. Although water radiolysis is essential for radiation damage, all previous XPS studies were performed in vacuum. Here we present near-ambient-pressure XPS experiments to directly measure DNA damage under water atmosphere. They permit in-situ monitoring of the effects of radicals on fully hydrated double-stranded DNA. Our results allow us to distinguish direct damage, by photons and secondary low-energy electrons (LEE), from damage by hydroxyl radicals or hydration induced modifications of damage pathways. The exposure of dry DNA to x-rays leads to strand-breaks at the sugar-phosphate backbone, while deoxyribose and nucleobases are less affected. In contrast, a strong increase of DNA damage is observed in water, where OH-radicals are produced. In consequence, base damage and base release become predominant, even though the number of strand-breaks increases further. Furthermore, first data about the degradation of single-stranded DNA binding-proteins (G5P / GV5 and hmtSSB) under vacuum and NAP-XPS conditions are presented.
Radiation biophysics
(2023)
We give an overview about recent work concerning ionizing radiation damage to Oligonucleotides, plasmid DNA, DNA binding proteins (G5P), and DNA-protein complexes.
We focus on combining new experimental setups with Geant4/TOPAS particle scattering simulations to understand the effets of ionizing radiation.