Campus Units

Zoology, Molecular, Cellular and Developmental Biology, Genetics

Document Type

Article

Publication Version

Published Version

Publication Date

1995

Journal or Book Title

Biochemistry

Volume

34

Issue

14

First Page

4583

Last Page

4592

DOI

10.1021/bi00014a011

Abstract

Type I repeat sequences are evolutionarily conserved sequence elements found in the replication origin and transcriptional promoter region of the rRNA genes (rDNA) in Tetrahymena thermophila. An abundant single-stranded DNA binding protein, ssA-TIBF, specifically interacts with the A-rich strand of the Type I repeat sequence. Quantitative binding competition experiments performed with purified ssATIBF demonstrate that the binding site for ssA-TIBF includes sequences both within the conserved 33 nt element and in a 3' flanking region: addition of the 3' flanking sequence to the Type I repeat oligonucleotide increases the binding affinity of ssA-TIBF by nearly 100-fold (apparent K& = 3.0 x 10~10 M). A mutation in the ssA-TIBF binding site previously shown to be the determinant of an rDNA replication defect in vivo results in a 25-fold decrease in ssA-TIBF binding affinity in vitro. ssA-TIBF also binds with high affinity to a copy of the Type I repeat sequence within the essential promoter region defined by in vitro transcription assays. The affinity of ssA-TIBF for the promoter repeat, which differs from other copies of the repeat at 8 out of 33 positions, is at least equal to its affinity for the Type I repeat sequences in the origin region. The biochemical properties of ssA-TIBF in vitro suggest that it could play a role in both replication and transcription of Tetrahymena rDNA in vivo.

Comments

Reprinted (adapted) with permission from A single-stranded DNA binding protein that specifically recognizes cis-acting sequences in the replication origin and transcriptional promoter region of Tetrahymena rDNA, Zhen Hou, Angela R. Umthun, and Drena L. Dobbs, Biochemistry 1995 34 (14), 4583-4592, DOI: 10.1021/bi00014a011. Copyright 1995 American Chemical Society.

Copyright Owner

American Chemical Society

Language

en

File Format

application/pdf

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