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A single-stranded DNA binding protein that specifically recognizes cis-acting sequences in the replication origin and transcriptional promoter region of Tetrahymena rDNA.

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 ssA-TIBF 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 Kd = 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.

journal_name

Biochemistry

journal_title

Biochemistry

authors

Hou Z,Umthun AR,Dobbs DL

doi

10.1021/bi00014a011

subject

Has Abstract

pub_date

1995-04-11 00:00:00

pages

4583-92

issue

14

eissn

0006-2960

issn

1520-4995

journal_volume

34

pub_type

杂志文章

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