Exploration of the quantitative aspects of the MALDI TOFMS analysis of peptides

Andrew David Mahan

doi:10.17918/etd-4068

The focus of this thesis was on the analysis of peptides, paying particular attention to the physicochemical properties of the peptide which have the largest impact on the measured MALDI TOFMS signal. In the first project, typical MALDI samples containing matrix and analyte are investigated using electrospray ionization (ESI)-MS. The goal was to ascertain the role that the nonvolatile acidic matrix compound such as [alpha]-cyano-4-hydroxycinammic acid (CHCA) has on the observed charge states of peptides at increasing matrix-toanalyte ratios. In the second project, several methods of MALDI sample preparation are studied by MALDI Imaging to evaluate analyte homogeneity and signal reproducibility within a sample, as well as overall spectral quality. The third project involves a series of experiments designed to gain a better understanding of the effect of peptide structure on the MALDI TOFMS signal response. It is generally understood that increases in the hydrophobicity of the peptide leads to a decrease in the MALDI signal, while an increase in the number of basic groups present (lysine, arginine, histidine and the amine terminus) leads to large increases in the observed MALDI signal. This work goes a step further by investigating how the proximity of and exact location of multiple basic sites in the peptide chain affects the MALDI signal. The initial hypothesis was that increased separation of the basic sites would lead to the observation of higher signal by increasing the sphere of capture for protons in the desorbing MALDI plume. Rudimentary molecular modeling is used to rationalize the results obtained. The final project investigates the effect of the acid/base properties of both the peptide and matrix on the number of positive versus negative ions observed in the MALDI spectrum. This work explores the two-step model of ionization in MALDI described by Knockenmuss (Int. J. Mass Spectrom., 2008, 273, 84-86). In both cases electrospray deposition (rather than the conventional dried droplet method) is used to produce highly homogeneous MALDI samples that yield highly reproducible analyte ion signal. These results are important to those seeking to understand the observed sequence coverage obtained in peptide mass mapping or ladder sequencing type experiments.

Exploration of the quantitative aspects of the MALDI TOFMS analysis of peptides

Files and links (1)

Abstract

Metrics

Details

Exploration of the quantitative aspects of the MALDI TOFMS analysis of peptides

Files and links (1)

Abstract

Metrics

Details

Drexel University Social media