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You searched for subject:(DNA Pol III). Showing records 1 – 3 of 3 total matches.

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Northeastern University

1. Benson, Ryan W. Structure function analysis of Escherichia coli pol IV.

Degree: PhD, Department of Biology, 2013, Northeastern University

Translesion synthesis (TLS) DNA polymerases are low fidelity DNA polymerases responsible for inserting a deoxynucleotide opposite to and extending from replication-blocking DNA lesions that have evaded high fidelity DNA repair pathways. E. coli DinB (DNA Pol IV) is a TLS polymerase of great interest because of its conservation throughout all domains of life and its relatively high intracellular basal level when compared to other DNA polymerases. Alternative activities of DinB other than its TLS ability include: roles in recombination, transcription, and a DNA replication "checkpoint." Using a structure/function strategy, we sought to better understand these activities by utilizing dinB alleles with different catalytic activities, in combination with other mutated genes involved in DNA replication or repair. This approach has allowed us to identify residues of DinB important for its various cellular tasks. In this work we have identified the roles of three highly conserved catalytic residues in the fidelity of lesion bypass of diverse lesions.; We also addressed the underlying mechanisms governing interactions between different bacterial DNA polymerases upon DNA damage treatment, which is likely to activate either DNA polymerase exchange and/or DNA replication checkpoints. By using the dnaE915 allele, encoding a variant of the replicative DNA Pol III catalytic α-subunit (Pol IIIα), we detected a DNA damage treatment-dependent growth arrest at in vivo DinB intracellular concentrations much lower than those in previous studies. A variety of intragenic dinB mutations localized to a specific area of DinB that suppressed any observed growth defects were also identified. We infer that this DinB face is interacting directly or indirectly with DNA Pol IIIα.; Lastly, we discovered a novel TLS independent role of DinB in the tolerance of DNA damage treatment. However, this role does not simply involve protein-protein interactions but requires the catalytic activity of DinB. Therefore, survival is likely the result of the largely error free synthesis of undamaged DNA that is independent of recA mediated recombination, induction of the SOS DNA damage response, or base excision repair. The structure/function analysis undertaken in this work has allowed us to discern and explore several activities of DinB other than its part in canonical DNA damage tolerance through strict TLS. This information will allow greater understanding of the mechanisms regulating both the TLS activity and other alternative roles that these specialized DNA polymerases might be playing in cells experiencing DNA damage or environmental stress.

Subjects/Keywords: DinB; DNA Pol III; DNA Pol IV; E. coli; TLS; Biology

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APA (6th Edition):

Benson, R. W. (2013). Structure function analysis of Escherichia coli pol IV. (Doctoral Dissertation). Northeastern University. Retrieved from http://hdl.handle.net/2047/d20003117

Chicago Manual of Style (16th Edition):

Benson, Ryan W. “Structure function analysis of Escherichia coli pol IV.” 2013. Doctoral Dissertation, Northeastern University. Accessed June 17, 2019. http://hdl.handle.net/2047/d20003117.

MLA Handbook (7th Edition):

Benson, Ryan W. “Structure function analysis of Escherichia coli pol IV.” 2013. Web. 17 Jun 2019.

Vancouver:

Benson RW. Structure function analysis of Escherichia coli pol IV. [Internet] [Doctoral dissertation]. Northeastern University; 2013. [cited 2019 Jun 17]. Available from: http://hdl.handle.net/2047/d20003117.

Council of Science Editors:

Benson RW. Structure function analysis of Escherichia coli pol IV. [Doctoral Dissertation]. Northeastern University; 2013. Available from: http://hdl.handle.net/2047/d20003117


Northeastern University

2. Sharma, Rajal. Engineering Escherichia coli DNA Polymerase III α for translesion synthesis.

Degree: MS, Department of Chemistry and Chemical Biology, 2010, Northeastern University

Accurate DNA replication is paramount for survival of all organisms. High-fidelity DNA polymerases ensure correct geometry of newly-forming base pairs by utilizing a tight fitting active site that allows only the correct incoming nucleotide to bind. This is reflected in the low error rate of high-fidelity polymerases; only 1 error in over 103 to 105 nucleotides incorporated. However this high geometric specificity can be disrupted. DNA is constantly under assault by mutagens, carcinogens, and reactive metabolic products. These agents can form adducts on DNA that disrupt high-fidelity DNA polymerases and stall the replication fork. The newly discovered Y family of DNA polymerases can bypass these lesions and allow the replication fork to continue. I am interested in the factors that make the major replicative DNA polymerase in E. coli, the DNA pol III alpha subunit, a high-fidelity polymerase. We have built a homology model of the enzyme in complex with DNA. We have identified potential residues important for fidelity. These residues were mutated and the polymerase variants were assayed by primer extension analysis on damaged and undamaged DNA. Engineering a low-fidelity polymerase from a high-fidelity polymerase allows us to identify the factors controlling polymerase specificity. Our computational models and kinetic data help to reveal the factors important in replication of damaged DNA.

Subjects/Keywords: chemistry; biochemistry; DNA Damage; DNA replication; dnaE; Pol III; polymerase; DNA adducts; DNA replication; DNA polymerases; Escherichia coli; Biochemistry; Cell Biology

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APA · Chicago · MLA · Vancouver · CSE | Export to Zotero / EndNote / Reference Manager

APA (6th Edition):

Sharma, R. (2010). Engineering Escherichia coli DNA Polymerase III α for translesion synthesis. (Masters Thesis). Northeastern University. Retrieved from http://hdl.handle.net/2047/d20000331

Chicago Manual of Style (16th Edition):

Sharma, Rajal. “Engineering Escherichia coli DNA Polymerase III α for translesion synthesis.” 2010. Masters Thesis, Northeastern University. Accessed June 17, 2019. http://hdl.handle.net/2047/d20000331.

MLA Handbook (7th Edition):

Sharma, Rajal. “Engineering Escherichia coli DNA Polymerase III α for translesion synthesis.” 2010. Web. 17 Jun 2019.

Vancouver:

Sharma R. Engineering Escherichia coli DNA Polymerase III α for translesion synthesis. [Internet] [Masters thesis]. Northeastern University; 2010. [cited 2019 Jun 17]. Available from: http://hdl.handle.net/2047/d20000331.

Council of Science Editors:

Sharma R. Engineering Escherichia coli DNA Polymerase III α for translesion synthesis. [Masters Thesis]. Northeastern University; 2010. Available from: http://hdl.handle.net/2047/d20000331


University of California – San Diego

3. KANG, JIN JOO. Mapping the Interactions of the Multi-Subunit General Transcription Factor IIIB (TFIIIB) with U6 Promoter DNA by Site-Specific Protein-DNA Photo-Cross-Linking.

Degree: Chemistry (Joint Doctoral SDSU), 2016, University of California – San Diego

U6 small nuclear RNA (snRNA) functions as a component of the spliceosome and is required to splice eukaryotic pre-mRNAs. Although the other spliceosomal snRNAs are synthesized by RNA polymerase II, U6 snRNA is synthesized by RNA polymerase III. The genes that code for U6 snRNA have promoters that are completely gene-external, that is, in the 5'-flanking DNA. These promoters contain a TATA box, as well as a required upstream proximal sequence element (PSE) that is unique to snRNA genes. The PSE is recognized by a multi-subunit transcription factor SNAPc, whereas the TATA sequence is bound by the general transcription factor TFIIIB, which is responsible for assembling an RNA polymerase III initiation complex. In the fruit fly Drosophila melanogaster, TFIIIB is composed of three distinct subunits that assemble on the U6 promoter: TBP, Bdp1 and Brf1. [On tRNA and 5S rRNA promoters in fruit flies, TRF1 (TBP-related factor 1) is employed in place of TBP.] At the present time, the overall structures of TFIIIB and particularly the Brf1 and Bdp1 subunits are not well known. To better understand the mode of fruit fly TFIIIB binding to DNA, I have used site-specific protein-DNA photo-cross-linking. I introduced photo-cross-linker at every second phosphate position in the DNA backbone on both the template and non-template strands within and surrounding the fly U6 TATA box. I have in this manner mapped the locations where each of the TFIIIB subunits lies in close proximity to the DNA. Brf1 cross-links strongly to the DNA within and upstream of the TATA box, whereas Bdp1 cross-links strongly both upstream and downstream but not within the TATA sequence. The data indicate that Bdp1 lies on the DNA in close proximity to two subunits of DmSNAPc. Bdp1 may therefore play a primary role in the recruitment of TFIIIB to the U6 promoter through interactions with DmSNAPc.

Subjects/Keywords: Biochemistry; Pol III; Site-specific Protein-DNA photo-Cross-Linking; TATA box; TFIIIB; U6 promoter; U6 snRNA

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APA · Chicago · MLA · Vancouver · CSE | Export to Zotero / EndNote / Reference Manager

APA (6th Edition):

KANG, J. J. (2016). Mapping the Interactions of the Multi-Subunit General Transcription Factor IIIB (TFIIIB) with U6 Promoter DNA by Site-Specific Protein-DNA Photo-Cross-Linking. (Thesis). University of California – San Diego. Retrieved from http://www.escholarship.org/uc/item/96p7v2xh

Note: this citation may be lacking information needed for this citation format:
Not specified: Masters Thesis or Doctoral Dissertation

Chicago Manual of Style (16th Edition):

KANG, JIN JOO. “Mapping the Interactions of the Multi-Subunit General Transcription Factor IIIB (TFIIIB) with U6 Promoter DNA by Site-Specific Protein-DNA Photo-Cross-Linking.” 2016. Thesis, University of California – San Diego. Accessed June 17, 2019. http://www.escholarship.org/uc/item/96p7v2xh.

Note: this citation may be lacking information needed for this citation format:
Not specified: Masters Thesis or Doctoral Dissertation

MLA Handbook (7th Edition):

KANG, JIN JOO. “Mapping the Interactions of the Multi-Subunit General Transcription Factor IIIB (TFIIIB) with U6 Promoter DNA by Site-Specific Protein-DNA Photo-Cross-Linking.” 2016. Web. 17 Jun 2019.

Vancouver:

KANG JJ. Mapping the Interactions of the Multi-Subunit General Transcription Factor IIIB (TFIIIB) with U6 Promoter DNA by Site-Specific Protein-DNA Photo-Cross-Linking. [Internet] [Thesis]. University of California – San Diego; 2016. [cited 2019 Jun 17]. Available from: http://www.escholarship.org/uc/item/96p7v2xh.

Note: this citation may be lacking information needed for this citation format:
Not specified: Masters Thesis or Doctoral Dissertation

Council of Science Editors:

KANG JJ. Mapping the Interactions of the Multi-Subunit General Transcription Factor IIIB (TFIIIB) with U6 Promoter DNA by Site-Specific Protein-DNA Photo-Cross-Linking. [Thesis]. University of California – San Diego; 2016. Available from: http://www.escholarship.org/uc/item/96p7v2xh

Note: this citation may be lacking information needed for this citation format:
Not specified: Masters Thesis or Doctoral Dissertation

.