Date of Award

January 2016

Degree Type

Thesis

Degree Name

Master of Science (MS)

Department

PULSe

First Advisor

Chittaranjan Das

Committee Member 1

Jean-Christophe Rochet

Committee Member 2

Mark Hall

Committee Member 3

Andrew Mesecar

Abstract

ElaD is a cysteine protease found in Escherichia coli (E. coli) and has been shown to function as a deubiquitinating enzyme (DUB). However, ubiquitin and the ubiquitination system are exclusive to eukaryotic cells. This indicates that ElaD may be used when the bacteria come in contact with a eukaryotic cell. This explains its presence in the intestinally infective strains of E. coli. For the invasive strains of E. coli, membrane fusion and phagosome formation is used for entry of the infective cell into the host cell. In order to counteract this, ubiquitination, the eukaryotic cell’s first defense mechanism, is used to signal for degradation of the phagosome. The phagosome is signaled for degradation by the host via adornment of the phagosome with Lys63-linked ubiquitin chains. We hypothesize that ElaD is used to neutralize ubiquitination of the phagosome and prevent destruction of the bacteria. With this hypothesis in mind we seek to examine the substrate preference of the DUB using diubiquitin substrates of defined linkage types. In addition we wish to understand the role of the active-site loop in determining selectivity of the DUB for ubiquitin over a ubiquitin-like modifier Nedd8.

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