CaltechTHESIS
  A Caltech Library Service

Bioorthogonal Noncanonical Amino Acid Tagging for Selective Analysis of the Pseudomonas aeruginosa Proteome

Citation

Babin, Brett Michael (2016) Bioorthogonal Noncanonical Amino Acid Tagging for Selective Analysis of the Pseudomonas aeruginosa Proteome. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/Z94X55S1. https://resolver.caltech.edu/CaltechTHESIS:05062016-131543245

Abstract

In natural environments, bacterial physiology is frequently characterized by slow metabolic rates and complex cellular heterogeneities. The opportunistic pathogen Pseudomonas aeruginosa provides one such example; P. aeruginosa forms untreatable chronic biofilm infections of the cystic fibrosis lung, where oxygen limitation can lead to states of metabolic dormancy. To better understand the biology of these states, in vitro experiments must be adapted to better recapitulate natural settings. However, low rates of protein turnover and cellular or phenotypic complexity make these systems difficult to study using established methods. Here we adapt the bioorthogonal noncanonical amino acid tagging (BONCAT) method for time- and cell-selective proteomic analysis to the study of P. aeruginosa. Analysis of proteins synthesized in an anoxic dormancy state led to the discovery of a new type of transcriptional regulator which we designated SutA. We performed detailed analyses of SutA’s role in transcription under slow growth states and we elucidated the structural basis for its regulatory behavior. Additionally, we used cell-selective targeting of BONCAT labeling to measure the dynamic proteomic response of an antibiotic-tolerant biofilm subpopulation. Overall this work shows the utility of selective proteomics as applied to bacterial physiology and describes the broad biological insight obtained from that application.

Item Type:Thesis (Dissertation (Ph.D.))
Subject Keywords:BONCAT;proteomics;Pseudomonas aeruginosa;transcription;biofilms
Degree Grantor:California Institute of Technology
Division:Chemistry and Chemical Engineering
Major Option:Chemical Engineering
Thesis Availability:Public (worldwide access)
Research Advisor(s):
  • Tirrell, David A.
Thesis Committee:
  • Tirrell, David A. (chair)
  • Newman, Dianne K.
  • Arnold, Frances H.
  • Ismagilov, Rustem F.
Defense Date:10 May 2016
Funders:
Funding AgencyGrant Number
U.S. Army Research OfficeW911NF-09-0001
Record Number:CaltechTHESIS:05062016-131543245
Persistent URL:https://resolver.caltech.edu/CaltechTHESIS:05062016-131543245
DOI:10.7907/Z94X55S1
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1021/cb300238wDOIState-Selective Metabolic Labeling of Cellular Proteins
http://dx.doi.org/10.1111/1462-2920.12436DOIIn situ visualization of newly synthesized proteins in environmental microbes using amino acid tagging and click chemistry
http://dx.doi.org/10.1073/pnas.1514412113DOISutA is a bacterial transcription factor expressed during slow growth in Pseudomonas aeruginosa
ORCID:
AuthorORCID
Babin, Brett Michael0000-0002-4133-6665
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:9704
Collection:CaltechTHESIS
Deposited By: Brett Babin
Deposited On:24 May 2016 21:59
Last Modified:04 Oct 2019 00:13

Thesis Files

[img]
Preview
PDF (Full thesis) - Final Version
See Usage Policy.

10MB
[img]
Preview
PDF (Thesis - Introduction and Chapter 1) - Final Version
See Usage Policy.

1MB
[img]
Preview
PDF (Thesis - Chapter 2) - Final Version
See Usage Policy.

11MB
[img]
Preview
PDF (Thesis - Chapter 3) - Final Version
See Usage Policy.

7MB
[img]
Preview
PDF (Thesis - Chapter 4) - Final Version
See Usage Policy.

6MB
[img]
Preview
PDF (Thesis - Appendices) - Final Version
See Usage Policy.

5MB
[img] MS Excel (Dataset_4_1) - Supplemental Material
See Usage Policy.

179kB
[img] MS Excel (Dataset_4_2) - Supplemental Material
See Usage Policy.

303kB
[img] MS Excel (Dataset_A_1) - Supplemental Material
See Usage Policy.

83kB
[img] MS Excel (Dataset_A_2) - Supplemental Material
See Usage Policy.

22kB
[img] MS Excel (Dataset_A_3) - Supplemental Material
See Usage Policy.

1MB
[img] MS Excel (Dataset_A_4) - Supplemental Material
See Usage Policy.

18kB
[img] MS Excel (Dataset_A_5) - Supplemental Material
See Usage Policy.

18MB
[img] MS Excel (Dataset_A_6) - Supplemental Material
See Usage Policy.

285kB

Repository Staff Only: item control page