| Mass spectrometry (MS) constitutes a powerful technique for studying the chemical modifications of proteins and DNA. This dissertation consists of several topics on using mass spectrometry to study protein glycation, peptide phosphorylation, and DNA crosslink and methylation.; Chapter 2 involved the systematic study of the methylglyoxal (MG)-induced modification of human hemoglobin. The treatment of human hemoglobin with MG in vitro could induce MG-H1 modification on arginine residues and the modification levels correlated well with the solvent accessibilities of those arginine residues.; In chapter 3, the MG-induced arginine modification of hemoglobin was further examined in diabetic mouse blood samples. Although LC-MS-based HbA 1c assay suggested that more severe glycemia occurred in diabetic mice with nephropathy than that in diabetic mice, no obvious correlation between the MG-H1 level and the HbA1c level was observed in diabetic mice blood either with or without nephropathy.; In chapter 4, the MG-induced modification of alpha-crystallin was assessed. Although the arginine MG-H1 modification could be unambiguously detected in bovine alpha-crystallin via treatment with MG in vitro, we were unable to observe the same modification in alpha-crystallin isolated from diabetic mouse lens.; In chapter 5, a simple method was introduced to measure the ionization efficiency changes of peptides introduced by their PTMs. We found that the ionization efficiency of alanine-rich peptides could be altered by threonine phosphorylation. In addition, the location of threonine phosphorylation could affect the ionization efficiency of the peptide.; In chapter 6, we examined the structures of a new DNA photoproduct formed between neighboring cytosine residues. We found that Pyrex-filtered UV light irradiation of d(BrCC) ("BrC" represents 5-bromocytosine) could give rise to three types of intrastrand crosslink products. Furthermore, we identified the structure of the most stable product, that is d(C[5-5]C), where the C5 carbon atom of 5' cytosine is covalently bonded to C5 carbon atom of 3' cytosine.; In chapter 7, the N6 methylation of adenine in different living organisms was quantitatively assessed by using LC-MS with the isotope-dilution method. We provided solid mass spectrometric evidence supporting the presence of N6-methyl-2'-deoxyadenosine in the Saccharomyces cerevisiae and Arabidopsis thaliana . |
