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The Bloom's syndrome DNA helicase complex
Descriptive
TypeOfResource
Text
TitleInfo
(ID = T-1)
Title
The Bloom's syndrome DNA helicase complex
SubTitle
identification and characterization of activities conserved in the orthologous complex from saccharomyces cerevisiae
Identifier
ETD_2579
Identifier
(type = hdl)
http://hdl.rutgers.edu/1782.1/rucore10001600001.ETD.000053567
Language
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(type = code)
eng
Genre
(authority = marcgt)
theses
Subject
(ID = SBJ-1);
(authority = RUETD)
Topic
Biochemistry
Subject
(ID = SBJ-2);
(authority = ETD-LCSH)
Topic
Genomics
Subject
(ID = SBJ-3);
(authority = ETD-LCSH)
Topic
DNA helicases
Abstract
(type = abstract)
Bloom's Syndrome (BS) is a rare human disease characterized by genome instability and cancer predispostion. The gene mutated in BS, BLM, encodes a member of the RecQ family of DNA helicases. This family consists of five human paralogs that play crucial roles in guarding against DNA rearrangements. All BLM orthologs, including budding yeast Sgs1, bind stably to a protein complex composed of DNA topoisomerase 3α (Top3) and the OB-fold protein Rmi1. Although the BLM/Sgs1 complex is known to suppress homologous recombination, it's mechanism of action is unknown. I found that a stable Top3-Rmi1 complex can be isolated from yeast cells overexpressing these two subunits and it shows increased superhelical relaxation activity compared to Top3 alone. The Rmi1 subunit also stimulates Top3 activity in reconstitution experiments. In both cases, elevated temperatures are required for optimal relaxation unless the substrate contains a ssDNA bubble. Interestingly, Rmi1 binds only weakly to ssDNA on its own, but it stimulates the ssDNA binding activity of Top3 five-fold. Top3 and Rmi1 also cooperate to bind the Sgs1 N-terminus and promote its interaction with single-strand (ss) DNA. In addition to the highly-conserved DNA helicase domain, all BLM/Sgs1 orthologs contain a large (652 aa) N-terminal domain that has no known catalytic activity. To determine the function of the N-terminal domain, I assayed truncated Sgs1 proteins for ssDNA binding activity. I identified a sub-domain of the Sgs1 N-terminus (SE, aa #103-322) that displays in vitro ssDNA binding, ssDNA annealing and strand exchange (SE) activities. These activities are conserved in the human and Drosophila orthologs. Strand exchange between duplex DNA and homologous ssDNA requires no cofactors and is inhibited by a single mismatched base-pair. The SE domain of Sgs1 is required in vivo for the suppression of hyper-recombination, suppression of synthetic-lethality and heteroduplex rejection. The top3∆ slow-growth phenotype is also SE-dependent. Surprisingly, the highly divergent SE domain from human BLM functions in yeast. Thus, SE activity is a new molecular function of BLM/Sgs1 that is conserved in other recombinases. The data suggest that at least one role of SE is to mediate the strand-passage events catalyzed by Top3-Rmi1.
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electronic resource
Extent
ix, 139 p. : ill.
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application/pdf
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Note
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Ph.D.
Note
Includes abstract
Note
Vita
Note
(type = bibliography)
Includes bibliographical references
Note
(type = statement of responsibility)
by Chi-Fu Chen
Name
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Chen
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Chi-Fu
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1975-
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author
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Chi-Fu Chen
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Brill
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Steven J.
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chair
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Advisory Committee
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Steven J. Brill
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Brenneman
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Mark
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internal member
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Advisory Committee
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Mark Brenneman
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Gartenberg
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Marc
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internal member
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Advisory Committee
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Marc Gartenberg
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Walworth
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Nancy C.
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outside member
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Advisory Committee
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Nancy C. Walworth
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Rutgers University
Role
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degree grantor
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Graduate School - New Brunswick
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school
OriginInfo
DateCreated
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2010
DateOther
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2010
Place
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xx
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TitleInfo
Title
Rutgers University Electronic Theses and Dissertations
Identifier
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ETD
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Title
Graduate School - New Brunswick Electronic Theses and Dissertations
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rucore19991600001
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(displayLabel = Rutgers, The State University of New Jersey)
NjNbRU
Identifier
(type = doi)
doi:10.7282/T3GF0TKG
Genre
(authority = ExL-Esploro)
ETD doctoral
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Rights
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The author owns the copyright to this work.
Copyright
Status
Copyright protected
Availability
Status
Open
Reason
Permission or license
RightsHolder
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(type = personal)
Name
FamilyName
Chen
GivenName
Chi-Fu
Role
Copyright Holder
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Type
Permission or license
DateTime
2010-04-13 13:42:07
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Name
Chi-Fu Chen
Affiliation
Rutgers University. Graduate School - New Brunswick
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Author Agreement License
Detail
I hereby grant to the Rutgers University Libraries and to my school the non-exclusive right to archive, reproduce and distribute my thesis or dissertation, in whole or in part, and/or my abstract, in whole or in part, in and from an electronic format, subject to the release date subsequently stipulated in this submittal form and approved by my school. I represent and stipulate that the thesis or dissertation and its abstract are my original work, that they do not infringe or violate any rights of others, and that I make these grants as the sole owner of the rights to my thesis or dissertation and its abstract. I represent that I have obtained written permissions, when necessary, from the owner(s) of each third party copyrighted matter to be included in my thesis or dissertation and will supply copies of such upon request by my school. I acknowledge that RU ETD and my school will not distribute my thesis or dissertation or its abstract if, in their reasonable judgment, they believe all such rights have not been secured. I acknowledge that I retain ownership rights to the copyright of my work. I also retain the right to use all or part of this thesis or dissertation in future works, such as articles or books.
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ETD
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application/pdf
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application/x-tar
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16148480
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