The functional subunit of a dimeric transcription activator protein depends on promoter architecture.

Zhou, Yuhong; Pendergrast, P. S.; Bell, Andrew I.; Williams, Roy M.; Busby, Stephen; Ebright, Richard H.

doi:10.1002/j.1460-2075.1994.tb06776.x

Cited by 90 publications

(62 citation statements)

References 40 publications

(45 reference statements)

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“…Therefore, a different set of side chains, but probably the same region of ␣-CTD, might be important for activation by CRP and CooA. This hypothesis is consistent with the low similarity between CRP and CooA in activating region 1 (22). In the DNase I footprinting analysis, RNAP does not completely protect the DNA downstream of P cooF ϩ1, even in the presence of CooA and in the absence of heparin (Fig.…”

Section: Discussionsupporting

confidence: 71%

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Transcription Activation by CooA, the CO-sensing Factor fromRhodospirillum rubrum

Gaál

Karls

et al. 1999

Journal of Biological Chemistry

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CooA, a member of the cAMP receptor protein (CRP) family, is a CO-sensing transcription activator from Rhodospirillum rubrum that binds specific DNA sequences in response to CO. The location of the CooAbinding sites relative to the start sites of transcription suggested that the CooA-dependent promoters are analogous to class II CRP-dependent promoters. In this study, we developed an in vivo CooA reporter system in Escherichia coli and an in vitro transcription assay using RNA polymerases (RNAP) from E. coli and from Rhodobacter sphaeroides to study the transcription properties of CooA and the protein-protein interaction between CooA and RNAP. The ability of CooA to activate CO-dependent transcription in vivo in heterologous backgrounds suggested that CooA is sufficient to direct RNAP to initiate transcription and that no other factors are required. This hypothesis was confirmed in vitro with purified CooA and purified RNAP. Use of a mutant form of E. coli RNAP with ␣ subunits lacking their Cterminal domain (␣-CTD) dramatically decreased CooAdependent transcription of the CooA-regulated R. rubrum promoter P cooF in vitro, which indicates that ␣-CTD plays an important role in this activation. DNase I footprinting analysis showed that CooA facilitates binding of wild-type RNAP, but not ␣-CTD-truncated RNAP, to P cooF . This facilitated binding provides evidence for a direct contact between CooA and ␣-CTD of RNAP during activation of transcription. Mapping the CooA-contact site in ␣-CTD suggests that CooA is similar but not identical to CRP in terms of its contact sites to the ␣-CTD at class II promoters.CooA is a CO-sensing transcription activator from Rhodospirillum rubrum, and it activates the expression of the cooFSCTJ and cooMKLXUH operons in response to CO (1-3). These two adjacent operons encode proteins that oxidize CO to CO 2 with concomitant reduction of H ϩ to H 2 and allow growth of this organism on CO as a sole energy source (4 -6). cooA lies immediately 3Ј of the cooFSCTJ operon, and its expression does not depend on CO.

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Section: Discussionsupporting

confidence: 71%

“…One interaction is between activating region 1 (AR1) of the upstream subunit of the CRP dimer and the ␣-CTD. This interaction increases initial binding of RNAP to the promoter (22). Recently, residues 285-288 and 317 of ␣-CTD have been shown to comprise the surface that interacts with AR1 of CRP at class II promoters (23).…”

mentioning

confidence: 99%

Transcription Activation by CooA, the CO-sensing Factor fromRhodospirillum rubrum

Gaál

Karls

et al. 1999

Journal of Biological Chemistry

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“…One possible rationale for this behavior is the requirement of recognition between two asymmetric macromolecules, namely CRP-cAMP 1 and CRP-dependent promoter, as proposed by Heyduk and Lee (17). The natural DNA sequences of CRP binding sites are not palindromic (1, 3), and RNA polymerase seems to recognize a DNAbound CRP subunit in a particular orientation (42,43). Hence, it is conceivable that CRP binds to the asymmetric DNA site in a specific orientation.…”

Section: Discussionmentioning

confidence: 99%

Interactive and Dominant Effects of Residues 128 and 141 on Cyclic Nucleotide and DNA Bindings in Escherichia coli cAMP Receptor Protein

Cheng¹,

Lee

1998

Journal of Biological Chemistry

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The molecular events in the cAMP-induced allosteric activation of cAMP receptor protein (CRP) involve interfacial communications between subunits and domains. However, the roles of intersubunit and interdomain interactions in defining the selectivity of cAMP against other cyclic nucleotides and cooperativity in ligand binding are still not known. Natural occurring CRP mutants with different phenotypes were employed to address these issues. Thermodynamic analyses of subunit association, protein stability, and cAMP and DNA binding as well as conformational studies of the mutants and wild-type CRPs lead to an identification of the apparently dominant roles of residues 128 and 141 in the cAMP-modulated DNA binding activity of CRP. Serine 128 and the C-helix were implicated as playing a critical role in modulating negative cooperativity of cyclic nucleotide binding. A correlation was established between a weak affinity for subunit assembly and the relaxation of cyclic nucleotide selectivity in the G141Q and S128A/ G141Q mutants. These results imply that intersubunit interaction is important for cyclic nucleotide discrimination in CRP. The double mutant S128A/G141Q, constructed from two single mutations of S128A and G141Q, which exhibit opposite phenotypic characteristics of CRP ؊ and CRP * , respectively, assumes a CRP * phenotype and has biochemical properties similar to those of the G141Q mutant. These observations suggest that mutation G141Q exerts a dominant effect over mutation S128A and that the subunit realignment induced by the G141Q mutation can override the local structural disruption created by mutation S128A.

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“…At some promoters, CRP binds upstream of the −35 promoter region, whereas at other promoters, the CRP site overlaps the −35 region. In the former case the promoter-proximal monomer of the CRP dimer is responsible for activation whereas in the latter case, the promoter-distal monomer activates transcription (Zhou et al, 1993a(Zhou et al, , 1994b. Thus, which CRP monomer is functional in activation depends on the position of the binding site with respect to the promoter.…”

Section: Discussionmentioning

confidence: 99%

Role of Operator Subsites in Arc Repression

Smith

Sauer

1996

Journal of Molecular Biology

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The functional subunit of a dimeric transcription activator protein depends on promoter architecture.

Cited by 90 publications

References 40 publications

Transcription Activation by CooA, the CO-sensing Factor fromRhodospirillum rubrum

Transcription Activation by CooA, the CO-sensing Factor fromRhodospirillum rubrum

Interactive and Dominant Effects of Residues 128 and 141 on Cyclic Nucleotide and DNA Bindings in Escherichia coli cAMP Receptor Protein

Role of Operator Subsites in Arc Repression

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