Date of Award

1-1-2013

Language

English

Document Type

Master's Thesis

Degree Name

Master of Science (MS)

College/School/Department

Department of Chemistry

Content Description

1 online resource (x, 80 pages) : illustrations (some color)

Dissertation/Thesis Chair

Carla Theimer

Keywords

pseudoknot, RNA, telomerase, Telomere, Telomerase, Nucleosides

Subject Categories

Biochemistry

Abstract

Telomerase is a ribonucleoprotein (RNP) complex that synthesizes telomeric repeats at the ends of linear chromosomes to form the DNA–protein complexes known as telomeres. Telomeres protect the ends of chromosomes from degradation during replication due to the end-replication problem. When replication occurs, gaps are created at the beginning of the lagging and leading strands that result in the loss of a small amount of DNA at every replication cycle. By adding thousands of copies of telomeric repeats, the repeats are lost in the replication process and not precious genetic information. The telomerase RNA varies drastically among different species both in size and sequence, but maintains the common core structural motif of an RNA pseudoknot. The catalytic function of the pseudoknot was tested using 2’– O–methyl substitutions in the pseudoknot stem 2 and tertiary structure. 2’–O–methyl substitutions were chosen because they were expected to be thermodynamically and structurally conservative. Circular dichroism (CD) spectroscopy was used to determine the effects of these substitutions on the structure of the pseudoknot. To ensure all pseudoknot complexes were made in a 1:1 ratio with no excess RNA, native polyacrylamide gel electrophoresis was used to separate the CD samples by size and compare them to standards. Once the complexes were verified, thermal melting profiles of the complexes were measured to determine the stabilities of the bimolecular pseudoknots, for comparison with catalytic activity. This research leads the way for further study of the telomerase pseudoknot and how small substitutions can affect telomerase activity.

Download

Included in

Biochemistry Commons

Share

COinS