Study of oxidation of thiols and disulfide bonds by singlet oxygen and redox proteomics of keratinocytes exposed to light

• Grant number: 22/14455-9
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)
• Start date: November 01, 2023
• End date: July 31, 2027
• Field of knowledge: Biological Sciences - Biophysics - Radiology and Photobiology
• Principal Investigator: Paolo Di Mascio
• Grantee: Stella Boutris Jayme
• Host Institution: Instituto de Química (IQ). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

Singlet molecular oxygen (1O2) is a reactive oxygen species that, once present in cells, has the ability to oxidize biomolecules such as proteins, lipids and DNA. Previous results have shown that the reaction of 1O2 with thiols and disulfide bonds generates oxidation products in peptides and proteins. In light of this, the following research project seeks to study the mechanisms of oxidation in thiols and disulfide bridges by 1O2 in cysteine and cystine, and in peptides and proteins containing these amino acids, detecting the modifications by liquid chromatography coupled to mass spectrometry and isotopic labeling. After characterizing the thiol oxidation products in peptides and proteins, we will evaluate the effect of 1O2 oxidation in cells exposed to UVA and visible radiation through global redox proteomics. To test this hypothesis, cells of HaCaT lineage and NHEK primary keratinocytes will be used, irradiated with UVA and visible light and labeling of modifications in cysteines and disulfide bonds with N-ethylmaleimide or Iodine-TMT for analysis by mass spectrometry. The use of keratinocytes exposed directly to UVA and visible light seeks to verify the signature of oxidative damage in a biological model of solar radiation on the skin and to better understand the consequences of oxidation by 1O2. In this way, the present project aims to contribute to the understanding of the pathologies caused by this oxidant, especially those triggered by the modification in thiols and disulfide bonds. (AU)