Chemically Enhanced Fragmentation of Peptides Through Derivatization with N-Hydroxysuccinimidyl Ester of 3-Sulfobenzoic Acid

Reduced spectral complexity is an advantage offered by MALDI mass spectrometry due to its ability to generate almost exclusively singly-charged ions. However, this may limit the extent of fragmentation of peptides under CID conditions; due to the higher onset energy of fragmentation, when compared to the values for the multiply charged ions generated by electrospray ionization. Through modifying the N-terminal residue with a structural moiety featuring a sulfonic acid group, a second mobile proton can be incorporated into the peptide capable of enhancing fragmentation under collision-induced dissociation and post-source decay conditions.


Figure 1. Preparation of the derivatizing reagent. 3-sulfobenzoic acid is reacted with N-hydroxysuccinimide in the presence of dicyclohexylcarbodiimide and diisopropylethylamine in dimethylsulfoxide to produce this reagent.



Figure 2. MSMS Spectra for (upper trace) non-slufonated and (lower trace) sulfonated KVPQVSTPTLVEVSR peptide derived from bovine serum albumin.


Analytical/Proteomic Approach to Evaluating the Effect of Alcohol Intake on Liver Protein Expression and Modifications

Animal model is very invaluable for elucidating human nervous system, behavior or metabolic pathways in human organism; however, this model becomes useful when it can show relevant analogy about human conditions. For alcohol study is mostly used rat model with criteria for animal model [1].

Animal tissues were obtained from adult male rats (90-110 days old). P and iP line were developed and maintained at Indiana University School of Medicine. The animals were killed by decapitation and liver tissue was frozen immediately.

Inbred alcohol preferring rats (iP rats) were derived from a colony of Wistar rats. These rats are defined as animals with ethanol intakes of >= than 5 g/kg body weight /day. The iP line of rats is a well-established animal model of alcoholism. The aim of this study has been to determine the qualitative and quantitative effects of alcohol intake on protein expression in liver. The selectively bred high-preference animals received 15% ethanol for 8 weeks, while control group received only water. The total liver proteomes of two groups of rats were evaluated. The first group consisted of six individuals which are defined as ethanol-exposed rats receiving free access to 15% ethanol for 8 weeks, while another six individuals are defined as ethanol-naive rats, which received only water. The second group consists of the same number of animals as the first group.