Reaching Out Locating and Lengthening the Interdomain Linker in AraC Protein

Eustance, Rebecca J.; Bustos, Silvia A.; Schleif, Robert

doi:10.1016/s0022-2836(84)71584-1

Cited by 35 publications

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“…Our new results are somewhat surprising, in light of the earlier work on AraC (7,9). Previously, it was found that the interdomain linker was relatively tolerant to amino acid substitutions.…”

Section: Discussioncontrasting

confidence: 73%

“…Each monomer of the dimeric AraC protein (35) contains two structurally and functionally separate and distinct domains ( Fig. 1): a dimerization domain (DD) that both dimerizes the protein and binds arabinose and a DNA-binding domain (DBD) (1,9,27) that both binds to DNA and provides the interactions with RNA polymerase that stimulate transcription initiation at the ara p BAD promoter (40). In vivo in the absence of arabinose, AraC binds in trans to two well-separated DNA half-sites, araO 2 (O 2 ) and araI 1 (I 1 ) the binding represses p BAD activity by forming a DNA loop in which p BAD is inactive (4,16,18,19,21).…”

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confidence: 99%

“…Preliminary mutational studies of the linker of AraC were performed (7) once the approximate domain boundaries in AraC had been established with genetic studies (1,9). These mutational studies suggested that changes of a single amino acid in the linker produced minor effects but that changes of multiple residues might be capable of producing a substantial effect.…”

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confidence: 99%

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Active Role of the Interdomain Linker of AraC

Seedorff

Schleif

2011

Journal of Bacteriology

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The classical genetic studies of Englesberg and collaborators (5,6,32) showed that the product of the Escherichia coli araC gene positively and negatively regulates expression of the araBAD genes in response to the presence of arabinose. Subsequent biochemical and biophysical studies have provided in vitro assays for the protein's activities and have provided much information about the structure, properties, and function of the protein (reviewed in references 28 and 29). Each monomer of the dimeric AraC protein (35) contains two structurally and functionally separate and distinct domains (Fig.

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Section: Discussioncontrasting

confidence: 73%

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confidence: 99%

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Active Role of the Interdomain Linker of AraC

Seedorff

Schleif

2011

Journal of Bacteriology

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“…The N-terminal domain responds to effector and the C-terminal domain binds to site-specific DNA and activates transcription (6,10). The C-terminal domain of the AraC family has a helix-turn-helix motif and a high degree of sequence homology, where the consensus sequence of a characteristic conserved motif is I-DIA---GF-S--YF---F----G-TPS--R (5, 11).…”

Section: Discussionmentioning

confidence: 99%

maoB, a gene that encodes a positive regulator of the monoamine oxidase gene (maoA) in Escherichia coli

et al. 1996

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The structural gene for copper-and topa quinone-containing monoamine oxidase (maoA) and an unknown amine oxidase gene have been located at 30.9 min on the Escherichia coli chromosome. Deletion analysis showed that the unknown gene was located within a 1.1-kb cloned fragment adjacent to the maoA gene. The nucleotide sequence of this fragment was determined, and a single open reading frame (maoB) consisting of 903 bp was found. The gene encoded a polypeptide with a predicted molecular mass of 34,619 Da which was correlated with the migration on a sodium dodecyl sulfate-polyacrylamide gel. The predicted amino acid sequence of the MaoB protein was identical to the NH 2 -terminal amino acid sequence derived by Edman degradation of the protein synthesized under the self-promoter. No homology of the nucleotide sequence of maoB to the sequences of any reported genes was found. However, the amino acid sequence of MaoB showed a high level of homology with respect to the helix-turn-helix motif of the AraC family in its C terminus. The homology search and disruption of maoA on the chromosome led to the conclusion that MaoB is a transcriptional activator of maoA but not an amine oxidase. The consensus sequence of the cyclic AMP-cyclic AMP receptor protein complex binding domain was adjacent to the putative promoter for the maoB gene. By use of lac gene fusions with the maoA and maoB genes, we showed that the maoA gene is regulated by tyramine and MaoB and that the expression of the maoB gene is subject to catabolite repression. Thus, it seems likely that tyramine and the MaoB protein activate the transcription of maoA by binding to the regulatory region of the maoA gene.

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“…Based on this similarity, RhaS and RhaR are predicted to consist of two domains connected by a flexible linker (5,12,29,41). The N-terminal domains are predicted to be responsible for L-rhamnose binding and dimerization, while the C-terminal domains contain the 99-aminoacid region that classifies them as members of the AraC/XylS family.…”

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Amino Acid Contacts between Sigma 70 Domain 4 and the Transcription Activators RhaS and RhaR

Wickstrum

Egan

2004

J Bacteriol

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The RhaS and RhaR proteins are transcription activators that respond to the availability of L-rhamnose and activate transcription of the operons in the Escherichia coli L-rhamnose catabolic regulon. RhaR activates transcription of rhaSR, and RhaS activates transcription of the operon that encodes the L-rhamnose catabolic enzymes, rhaBAD, as well as the operon that encodes the L-rhamnose transport protein, rhaT. RhaS is 30% identical to RhaR at the amino acid level, and both are members of the AraC/XylS family of transcription activators. The RhaS and RhaR binding sites overlap the ؊35 hexamers of the promoters they regulate, suggesting they may contact the 70 subunit of RNA polymerase as part of their mechanisms of transcription activation. In support of this hypothesis, our lab previously identified an interaction between RhaS residue D241 and 70 residue R599. In the present study, we first identified two positively charged amino acids in 70 , K593 and R599, and three negatively charged amino acids in RhaR, D276, E284, and D285, that were important for RhaR-mediated transcription activation of the rhaSR operon. Using a genetic loss-of-contact approach we have obtained evidence for a specific contact between RhaR D276 and 70 R599. Finally, previous results from our lab separately showed that RhaS D250A and 70 K593A were defective at the rhaBAD promoter. Our genetic loss-of-contact analysis of these residues indicates that they identify a second site of contact between RhaS and 70 .Transcription activation in Escherichia coli often involves the interaction of a DNA-binding activator protein with one of the subunits of RNA polymerase (RNAP), most often the sigma () or alpha (␣) subunit. Transcription activators that bind immediately upstream and adjacent to RNAP, in some cases overlapping the Ϫ35 promoter hexamer, may interact with the C-terminal domain (domain 4) of the subunit of RNAP (8,27). The cI protein of bacteriophage is required for the establishment and maintenance of lysogeny and is perhaps the best-characterized example of a transcription activator that contacts 70 . The cI protein activates transcription of the P RM promoter when bound at the O R 2 operator site, which overlaps the P RM Ϫ35 hexamer by 2 bp (30). Current evidence suggests that 70 residues R588, K593, and R596 are required for activation by cI (23,26,35). Genetic and molecular modeling studies, as well as the recent structure of a ternary cI-domain 4-DNA complex, indicate that cI D38 contacts both 70 K593 ( A K418) and R596 ( A R421) and cI E34 contacts 70 R588 ( A R413) (8,19,26,35). Prior to the identification of the ternary complex structure, a molecular model of the interaction indicated that 70 K593 ( A K418) contacts DNA but was not positioned to contact cI (6,8,35). However, the ternary structure showed that the A residue that aligns with 70 K593 has moved away from the DNA (relative to the model of the interaction) and instead makes a proteinprotein contact with cI D38 (19).There is also evidence that activation by several tran...

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Reaching Out Locating and Lengthening the Interdomain Linker in AraC Protein

Cited by 35 publications

References 0 publications

Active Role of the Interdomain Linker of AraC

Active Role of the Interdomain Linker of AraC

maoB, a gene that encodes a positive regulator of the monoamine oxidase gene (maoA) in Escherichia coli

Amino Acid Contacts between Sigma 70 Domain 4 and the Transcription Activators RhaS and RhaR

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