Characterization of proteins and peptides via enhanced 266 nm ultraviolet photodissociation mass spectrometry utilizing a selenium based chromophore
| dc.contributor.advisor | Brodbelt, Jennifer S. | |
| dc.contributor.committeeMember | Eberlin, Livia S. | |
| dc.creator | Parker, William Ryan | |
| dc.creator.orcid | 0000-0001-5753-2508 | |
| dc.date.accessioned | 2018-08-17T18:37:24Z | |
| dc.date.available | 2018-08-17T18:37:24Z | |
| dc.date.created | 2016-05 | |
| dc.date.issued | 2016-05 | |
| dc.date.submitted | May 2016 | |
| dc.date.updated | 2018-08-17T18:37:24Z | |
| dc.description.abstract | Mass spectrometry and chemical derivatization have been used as tools for the identification of proteins in both top-down and bottom-up studies. Cysteine is the rarest and most nucleophilic amino acid thus making it a popular target for chemical tagging strategies. Ultraviolet photodissociation (UVPD) is a versatile activation technique for fragmentation of peptides and proteins. For successful photodissociation, ions of interest must contain a suitable chromophore that matches the wavelength of irradiation. N-(Phenylseleno)phthalimide (NPSP) is a fast reacting reagent which attaches a selenium based chromophore that absorbs at 266 nm light to free thiols. In the studies presented in this thesis, NPSP was used to derivatize free cysteine residues in both intact proteins and tryptic peptides. Activation with 266 nm photons causes a dominant neutral loss of the benzeneselenol groups on the tagged protein or peptide ions. This diagnostic neutral loss allows the determination of the number of free versus bound cysteine residues in intact proteins. Additionally, tagging peptides with benzeneselenol provides a means to target only the cysteine-containing peptides in bottom-up proteomics experiments. Both of these methods provide a significantly reduced search space for identification of cysteine-containing proteins. Counting the number of cysteine residues also provides an effective way to restrict the number of protein candidates for database searches. Moreover, cysteine peptides are inherently more unique than other peptides created upon enzymatic digestion of proteins due to the low frequency of cysteine in the proteome, thus allowing these peptides to be used as surrogates for protein identification. | |
| dc.description.department | Chemistry | |
| dc.format.mimetype | application/pdf | |
| dc.identifier | doi:10.15781/T2JW87613 | |
| dc.identifier.uri | http://hdl.handle.net/2152/68032 | |
| dc.language.iso | en | |
| dc.subject | Mass spectrometry | |
| dc.subject | 266 nm UVPD | |
| dc.subject | Proteomics | |
| dc.subject | Cysteine | |
| dc.title | Characterization of proteins and peptides via enhanced 266 nm ultraviolet photodissociation mass spectrometry utilizing a selenium based chromophore | |
| dc.type | Thesis | |
| dc.type.material | text | |
| thesis.degree.department | Chemistry | |
| thesis.degree.discipline | Chemistry | |
| thesis.degree.grantor | The University of Texas at Austin | |
| thesis.degree.level | Masters | |
| thesis.degree.name | Master of Arts |