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The S1 domains of Escherichia coli mRNA degradation Edge, Robert Edward
Abstract
The S1 domain is a recurring theme in the enzymes of mRNA decay in Escherichia coli, occurring in polynucleotide phosphorylase (PNPase), ribonuclease II (RNase II), ribonuclease E (RNase E) and ribonuclease G (RNase G). This thesis focuses on the function of the S1 domain of PNPase and RNase E. In the former, the hypothesis examined was that the domain provides an RNA -substrate contact that imparts processive phosphorylysis to the enzyme. Results show that PNPase lacking this domain still possesses catalytic activity, although at an extremely reduced rate, but appears to retain processive phosphorylysis of RNA substrates. In addition, the isolated PNPase domain has a Kd for RNA that is very high, suggesting that this domain may not play a role in the interactions between PNPase and its substrate. The S1 domain of RNase E was studied as an isolated domain. NMR experiments conducted by Dr. Mario Schubert revealed the solution structure of this isolated domain as well as potential dimerization contacts and identified residues implicated in nucleic acid interaction. Crosslinking data confirmed the dimerization of this domain. CD spectroscopy showed that a classical temperature -sensitive mutation in this domain alters its structure. The work on the RNase E S1 domain also tested the hypothesis that the domain confers the preference of RNase E for monophosphorylated substrates. The data suggest that this domain indeed binds to RNA (Kd 2 μM to 5 μM); however, it displays no increased affinity for differentially phosphorylated RNAs or detectable sequence specificity.
Item Metadata
Title |
The S1 domains of Escherichia coli mRNA degradation
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Creator | |
Publisher |
University of British Columbia
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Date Issued |
2003
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Description |
The S1 domain is a recurring theme in the enzymes of mRNA decay in Escherichia
coli, occurring in polynucleotide phosphorylase (PNPase), ribonuclease II (RNase II),
ribonuclease E (RNase E) and ribonuclease G (RNase G). This thesis focuses on
the function of the S1 domain of PNPase and RNase E. In the former, the
hypothesis examined was that the domain provides an RNA -substrate contact that
imparts processive phosphorylysis to the enzyme. Results show that PNPase
lacking this domain still possesses catalytic activity, although at an extremely
reduced rate, but appears to retain processive phosphorylysis of RNA substrates. In
addition, the isolated PNPase domain has a Kd for RNA that is very high, suggesting
that this domain may not play a role in the interactions between PNPase and its
substrate. The S1 domain of RNase E was studied as an isolated domain. NMR
experiments conducted by Dr. Mario Schubert revealed the solution structure of this
isolated domain as well as potential dimerization contacts and identified residues
implicated in nucleic acid interaction. Crosslinking data confirmed the dimerization of
this domain. CD spectroscopy showed that a classical temperature -sensitive
mutation in this domain alters its structure. The work on the RNase E S1 domain
also tested the hypothesis that the domain confers the preference of RNase E for
monophosphorylated substrates. The data suggest that this domain indeed binds to
RNA (Kd 2 μM to 5 μM); however, it displays no increased affinity for differentially
phosphorylated RNAs or detectable sequence specificity.
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Extent |
10854537 bytes
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Genre | |
Type | |
File Format |
application/pdf
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Language |
eng
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Date Available |
2009-11-17
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Provider |
Vancouver : University of British Columbia Library
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Rights |
For non-commercial purposes only, such as research, private study and education. Additional conditions apply, see Terms of Use https://open.library.ubc.ca/terms_of_use.
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DOI |
10.14288/1.0091167
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URI | |
Degree | |
Program | |
Affiliation | |
Degree Grantor |
University of British Columbia
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Graduation Date |
2003-11
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Campus | |
Scholarly Level |
Graduate
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Aggregated Source Repository |
DSpace
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Rights
For non-commercial purposes only, such as research, private study and education. Additional conditions apply, see Terms of Use https://open.library.ubc.ca/terms_of_use.