Peptidase activity of <i>Escherichia coli</i> aminopeptidase A is not required for its role in Xer site‐specific recombination

McCulloch, Richard; Burke, Mary Ellen; Sherratt, David J.

doi:10.1111/j.1365-2958.1994.tb01013.x

Cited by 65 publications

(58 citation statements)

References 35 publications

(22 reference statements)

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“…All plasmids, except those below, have been described Colloms et al, 1996). Plasmid pSH10 is a reporter plasmid containing two directly repeated cer sites flanking a lacZ gene; it is identical to pCS210 (McCulloch et al, 1994), except that the tetracyclineresistance (Tc R ) gene has been replaced by a kanamycinresistance (Km R ) gene. Plasmids pSH15-26 and pUC19-453 and pUC19-454, containing the 'idealized' ARG boxes and derivatives were derivatives of pUC19 containing the oligonucleotide sequences indicated in Fig.…”

Section: Bacteria and Plasmidsmentioning

confidence: 99%

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DNA binding of Escherichia coli arginine repressor mutants altered in oligomeric state

Chen

Merican

Sherratt

1997

Molecular Microbiology

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SummaryThe Escherichia coli arginine repressor (ArgR) controls expression of the arginine biosynthetic genes and acts as an accessory protein in Xer site-specific recombination at cer and related plasmid recombination sites. The hexameric wild-type protein shows L-arginine-dependent DNA binding. In this work, ArgR mutants that are defective in trimer-trimer interactions and bind DNA as trimers in an L-arginine-independent manner are isolated and characterized. Whereas the wild-type ArgR hexamer exhibits high-affinity binding to two repeated ARG boxes separated by 3 bp (each ARG box containing two identical dyad symmetrical 9 bp half-sites), the trimeric mutants bind to and footprint three adjacent half-sites of this 'idealized' substrate. Trimeric ArgR is impaired in its ability to repress the arginine biosynthetic genes and in Xer site-specific recombination. In the absence of L-arginine, residual wild-type ArgR-binding occurs as trimers. The binding of an N-terminal 77-amino-acid DNA-binding domain to idealized ARG boxes is also characterized.

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Section: Bacteria and Plasmidsmentioning

confidence: 99%

“…Assays of Xer site-specific recombination Intramolecular recombination at cer was measured by the loss of lacZ gene expression from the reporter plasmid pSH10, a relative of pCS210 (McCulloch et al, 1994;Burke et al, 1994), and by the loss of Cm R from pCS202 . Recombination in vitro was as in Colloms et al (1996).…”

Section: Gel Retardation Assaysmentioning

confidence: 99%

DNA binding of Escherichia coli arginine repressor mutants altered in oligomeric state

Chen

Merican

Sherratt

1997

Molecular Microbiology

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“…The E. coli LAP serves as an aminopeptidase (Vogt, 1970) and a DNA-binding protein that mediates both site-specific recombination at the cer site of ColE1 plasmids (Stirling et al, 1989) and transcriptional activation of the carAB operon (Charlier et al, 2000). The DNAbinding capabilities of the E. coli LAP are independent of aminopeptidase function (McCulloch et al, 1994;Charlier et al, 2000).…”

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confidence: 99%

Isolation and Characterization of the Neutral Leucine Aminopeptidase (LapN) of Tomato

Park

Walling

2003

Plant Physiology

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Tomatoes (Lycopersicon esculentum) express two forms of leucine aminopeptidase (LAP-A and LAP-N) and two LAP-like proteins. The relatedness of LAP-N and LAP-A was determined using affinity-purified antibodies to four LAP-A protein domains. Antibodies to epitopes in the most N-terminal region were able to discriminate between LAP-A and LAP-N, whereas antibodies recognizing central and COOH-terminal regions recognized both LAP polypeptides. Two-dimensional immunoblots showed that LAP-N and the LAP-like proteins were detected in all vegetative (leaves, stems, roots, and cotyledons) and reproductive (pistils, sepals, petals, stamens, and floral buds) organs examined, whereas LAP-A exhibited a distinct expression program. LapN was a single-copy gene encoding a rare-class transcript. A full-length LapN cDNA clone was isolated, and the deduced sequence had 77% peptide sequence identity with the wound-induced LAP-A. Leucyl aminopeptidases (LAPs; EC 3.4.11.1) are members of the M17 family of peptidases (Barrett et al., 1998). LAPs are ubiquitous being found in animals, plants, and prokaryotic cells. These hexameric metallopeptidases catalyze the release of the N-terminal residues from protein, peptide, fluorometric, and chromogenic substrates. The best characterized LAPs are from Bos taurus, Escherichia coli and tomato (Lycopersicon esculentum). X-ray crystal structures of the bovine and E. coli LAPs have provided insight into the LAP catalytic mechanism (Kim and Lipscomb, 1994;Sträter and Lipscomb, 1995;Sträter et al., 1999a). The roles of selected residues of the E. coli and tomato LAPs in chromogenic or peptide substrate catalysis, respectively, have been tested by site-directed mutagenesis (Sträter et al., 1999b;Gu and Walling, 2002).In most plants, three classes of LAP-related polypeptides are detected using a tomato LAP antiserum, including the 66-and 77-kD LAP-like proteins and the 55-kD neutral LAP (LAP-N; Chao et al., 2000). Only in a subset of the Solanaceae is a second 55-kD LAP species (LAP-A) detected (Hildmann et al., 1992;Gu et al., 1996b;Chao et al., 2000). In tomato, LAP-A protomers have an acidic pI and are encoded by two genes (LapA1 and LapA2), which are expressed during floral and fruit development. The LapA genes are not expressed in foliage from healthy plants (Chao et al., 1999). However, LapA RNAs, proteins, and activities accumulate locally and systemically in leaves after wounding, Pseudomonas syringae pv. tomato and Phytophthora parasitica infection, and caterpillar feeding (Pautot et al., 1993(Pautot et al., , 2001Gu et al., 1996b;Chao et al., 1999;Jwa and Walling, 2001). The activation of LapA gene expression by jasmonic acid (JA), abscisic acid, the phytotoxin coronatine (a JA mimic), and suppression of LapA by salicylic acid is consistent with the regulation of the tomato LapA genes by the wound octadecanoid pathway (Chao et al., 1999). LapA genes also respond to signals generated during water deficit and salinity stress (Chao et al., 1999). The potato (Solanum tuberosum) Lap RNAs also ac...

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“…PepA was expressed in DS941 arcA2 and purified as described by McCulloch et al (1994). His 6 -tagged ArcA was expressed in DS941 transformed with pQE30ArcA and pREP4.…”

Section: Proteinsmentioning

confidence: 99%

The ArcA/ArcB two‐component regulatory system of Escherichia coli is essential for Xer site‐specific recombination at psi

Colloms

Alén

Sherratt

1998

Molecular Microbiology

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SummaryTwo recombinases, XerC and XerD, act at the recombination sites psi and cer in plasmids pSC101 and ColE1 respectively. Recombination at these sites maintains the plasmids in a monomeric state and helps to promote stable plasmid inheritance. The accessory protein PepA acts at both psi and cer to ensure that only intramolecular recombination takes place. An additional accessory protein, ArgR, is required for recombination at cer but not at psi. Here, we demonstrate that the ArcA /ArcB two-component regulatory system of Escherichia coli, which mediates adaptation to anaerobic growth conditions, is required for efficient recombination in vivo at psi. Phosphorylated ArcA binds to psi in vitro and increases the efficiency of recombination at this site. Binding of ArcA to psi may contribute to the formation of a higher order synaptic complex between a pair of psi sites, thus helping to ensure that recombination is intramolecular.

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Peptidase activity of Escherichia coli aminopeptidase A is not required for its role in Xer site‐specific recombination

Cited by 65 publications

References 35 publications

DNA binding of Escherichia coli arginine repressor mutants altered in oligomeric state

DNA binding of Escherichia coli arginine repressor mutants altered in oligomeric state

Isolation and Characterization of the Neutral Leucine Aminopeptidase (LapN) of Tomato

The ArcA/ArcB two‐component regulatory system of Escherichia coli is essential for Xer site‐specific recombination at psi

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