Degree Type

Dissertation

Date of Award

1998

Degree Name

Doctor of Philosophy

Department

Chemistry

First Advisor

Cheng S. Lee

Second Advisor

Robert S. Houk

Abstract

On-line coupling of capillary isoelectric focusing (CIEF) and liquid chromatography (LC) with electrospray ionization mass spectrometry (ESIMS) is utilized for protein characterization;A microdialysis junction, based on a microdialysis membrane connecting both a separation capillary and a short, sharply tapered microelectrospray emitter capillary, is demonstrated for CIEF-ESIMS. Besides the electrical connection across the microdialysis junction, post-run acidification via the dialysis membrane is demonstrated for enhancing the protonation and the ionization efficiency of focused proteins while maintaining separation efficiency, resolution, and sensitivity in CIEF-ESIMS;On-line combination of CIEF with ESI-FTICRMS is demonstrated for high resolution analysis of model proteins, human hemoglobin variants, and Escherichia coli proteins. The acquisition of high-resolution mass spectra of hemoglobin [beta] chains allows direct identification of hemoglobin variants A and C, differing in molecular mass by 1 dalton. Direct mass determination of cellular proteins separated in the CIEF capillary is achieved using their isotopic envelopes obtained from ESI-FTICRMS;An integrated approach, involving the combination of exoglycosidase enzyme array with LC/MS/MS, is demonstrated for structural analysis of carbohydrate moieties of glycopeptides from a glycoprotein digest. The molecular mass information from a series of controlled digestions together with specific compositions of exoglycosidases in the enzyme array provide the sequence and linkage of individual glycan species attached to glycopeptides and glycoproteins.

DOI

https://doi.org/10.31274/rtd-180813-10751

Publisher

Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/

Copyright Owner

Liyu Yang

Language

en

Proquest ID

AAI9826588

File Format

application/pdf

File Size

109 pages

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