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Biochemical Characterization of hTRF1 and hTEP1, Two Proteins Involved in Telomere Maintenance

Tahmaseb, Kambiz
http://rave.ohiolink.edu/etdc/view?acc_num=wright1182306717

Abstract Details

2007, Doctor of Philosophy (PhD), Wright State University, Biomedical Sciences PhD.
Telomeres are the structures that protect the ends of linear chromosomes from fusion and degradation. The telomere consists of tandem repeated DNA sequences that can range from hundreds of bases to kilo-bases depending on the organism. As the cells of an organism replicate their DNA, these repeats are lost due to the end replication problem, where the ends of linear DNA cannot be fully replicated. As the telomeres are shortened through each round of replication, they eventually reach a critical point. Once the telomeres are too short and the cell risks losing coding sequences, a signaling pathway is initiated that causes the cell to senesce. However, cells that require continuous replication (i.e., stem cells, germ cells, and cancer cells) require constant maintenance of their telomeres in order to not enter senescence. The majority of these cells use the multimeric protein telomerase and a host of other proteins to maintain the lengths of their chromosomes. Eukaryotic telomerase is a nucleo-protein complex consisting of the telomerase RNA (TR), telomere end reverse transcriptase (TERT), and telomerase associated protein 1 (TEP1). Furthermore, telomeric length is regulated by a host of telomeric binding proteins. This thesis focuses on two proteins important for human telomeric maintenance. The first is human TEP1 (hTEP1) which is a subunit of telomerase. This large protein contains the RNA binding domain that binds hTR. Though the RNA binding subunit of hTEP1 has been partially purified before, full-length hTEP1 has been refractory to biochemical analysis due to the inability to express and purify this large protein. Here we reveal the very first purification of full-length hTEP1. Furthermore, where the RNA binding domain of hTEP1 alone does not show specific interaction with hTR, we show that full-length hTEP1 binds hTR specifically. The second protein of interest in this thesis is the human telomeric repeat binding factor 1 (hTRF1). This protein is one of the telomeric binding proteins that plays a critical role in telomere structure and stability. hTRF1 is also important as a regulator of telomeric length. hTRF1 has been shown to bind telomeric DNA specifically and my data reveals details of this surprisingly complex interaction using a sensitive intrinsic fluorescence kinetic technique. Our results demonstrate that hTRF1 binds to both telomeric and non-telomeric DNA. However, hTRF1 exhibits different characteristics as it binds telomeric DNA and is able to distinguish between telomeric and non-telomeric tracts of DNA. This new information on these two key players in the maintenance of telomeres will help us further understand how these complex DNA ends are preserved in the cell. Through this knowledge, we can devise better tests in understanding how immortal cell lines, such as cancer cells, function and proliferate, making it possible to identify novel therapeutic targets to inhibit this process.
John Turchi (Advisor)
122 p.
Chemistry, Biochemistry
Cancer; Fluorescence Quenching; Telomere; Telomerase; hTEP1; hTR; hTRF1

Recommended Citations

Citations

  • Tahmaseb, K. (2007). Biochemical Characterization of hTRF1 and hTEP1, Two Proteins Involved in Telomere Maintenance [Doctoral dissertation, Wright State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=wright1182306717

    APA Style (7th edition)

  • Tahmaseb, Kambiz. Biochemical Characterization of hTRF1 and hTEP1, Two Proteins Involved in Telomere Maintenance. 2007. Wright State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=wright1182306717.

    MLA Style (8th edition)

  • Tahmaseb, Kambiz. "Biochemical Characterization of hTRF1 and hTEP1, Two Proteins Involved in Telomere Maintenance." Doctoral dissertation, Wright State University, 2007. http://rave.ohiolink.edu/etdc/view?acc_num=wright1182306717

    Chicago Manual of Style (17th edition)

wright1182306717
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© 2007, all rights reserved.
This open access ETD is published by Wright State University and OhioLINK.