Relationship between promoter structure and template specificities exhibited by the bacteriophage T3 and T7 RNA polymerases.

Abstract:

:To explore the basis for the template specificities of the bacteriophage T3 and T7 RNA polymerases (EC 2.7.7.6), we determined the nucleotide sequences of six promoters recognized by the T3 RNA polymerase and compared them with the previously determined promoter sequences recognized by the bacteriophage T7 RNA polymerase. Recombinant plasmids containing random Hpa II and Taq I fragments of T3 DNA were screened for T3 promoter activity in vitro in a transcription assay using purified T3 RNA polymerase. Five promoters for the T3 RNA polymerase were identified in this manner and their sequences were determined; the sequence of an additional promoter was determined directly from a genomic DNA fragment. In five of the T3 promoters an identical 16-base-pair sequence (A-C-C-C-T-C-A-C-T-A-A-A-G-G-G-A) extends from -12 to +4 (initiation occurring with GTP at +1); this sequence is preceded by a 6-base-pair A + T region. The remaining promoter contains an inserted C at position -1 and an A at the +1 position. The sequence of the 5' end of the RNA transcript from the latter promoter confirms that transcription is initiated with ATP at the +1 position. Previously, late T3 or T7 transcripts had not been found to initiate with ATP. The highly conserved T3 promoter sequence was compared to the T7 promoter consensus sequence. The fundamental difference between the two kinds of phage promoters is the occurrence of G-A at positions -11 and -10 in the T7 promoter, whereas there is a single C at position -10 in the T3 promoter.

authors

Bailey JN,Klement JF,McAllister WT

doi

10.1073/pnas.80.10.2814

subject

Has Abstract

pub_date

1983-05-01 00:00:00

pages

2814-8

issue

10

eissn

0027-8424

issn

1091-6490

journal_volume

80

pub_type

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