Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.4.1 (phosphodiesterase)
18,767 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

During infection of homoimmune Escherichia coli lysogens ("repressed infections"), undamaged nonreplicating lambda phage DNA circles undergo very little recombination. Prior UV irradiation of phages dramatically elevates recombinant frequencies, even in bacteria deficient in UvrABC-mediated excision repair. We previously reported that 80-90% of this UvrABC-independent recombination required MutHLS function and unmethylated d(GATC) sites, two hallmarks of methyl-directed mismatch repair. We now find that deficiencies in other mismatch-repair activities--UvrD helicase, exonuclease I, exonuclease VII, RecJ exonuclease--drastically reduce recombination. These effects of exonuclease deficiencies on recombination are greater than previously observed effects on mispair-provoked excision in vitro. This suggests that the exonucleases also play other roles in generation and processing of recombinagenic DNA structures. Even though dsDNA breaks are thought to be highly recombinagenic, 60% of intracellular UV-irradiated phage DNA extracted from bacteria in which recombination is low--UvrD-, ExoI-, ExoVII-, or Rec(J-)--displays (near-)blunt-ended dsDNA ends (RecBCD-sensitive when deproteinized). In contrast, only bacteria showing high recombination (Mut+ UvrD+ Exo+) generate single-stranded regions in nonreplicating UV-irradiated DNA. Both recF and recB recC mutations strikingly reduce recombination (almost as much as a recF recB recC triple mutation), suggesting critical requirements for both RecF and RecBCD activity. The mismatch repair system may thus process UV-irradiated DNA so as to initiate more than one recombination pathway.
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PMID:DNA structures generated during recombination initiated by mismatch repair of UV-irradiated nonreplicating phage DNA in Escherichia coli: requirements for helicase, exonucleases, and RecF and RecBCD functions. 749 61

The single-stranded-DNA-binding (SSB) proteins from Proteus mirabilis and Serratia marcescens were purified from overproducing Escherichia coli strains, which were devoid of their own ssb gene. The strains harboured an endA insertion mutation and a xonA mutation resulting in the absence of endonuclease I and exonuclease I activities from the preparations. The amino acid sequences of the SSB of all three species are nearly identical in the N-terminal parts of the proteins that contain the DNA-binding domain, but differ in the C-terminal parts. Both proteins have an apparent binding-site size of 65 and 35 nucleotides at high and low salt concentrations, respectively. The association-rate constant for binding to poly(dT) is 3.2 x 10(8) M-1 s-1 for P. mirabilis SSB (PmiSSB) and 3.4 x 10(8) M-1 s-1 for S. marcescens SSB (SmaSSB). These binding parameters are very similar to those of E. coli SSB (EcoSSB). The structural similarity of the proteins is also documented by the finding that they can exchange subunits among each other to form mixed tetramers. The transcriptional regulation of the ssb and uvrA genes from P. mirabilis and S. marcescens in SOS-induced E. coli cells was studied using lacZ fusions. While the uvrA genes were inducible, there was no induction of the ssb genes transcribed divergently from the uvrA genes. Apparently, regions with nucleotide sequence similarity to the E. coli SOS-box preceding the ssb genes of P. mirabilis and S. marcescens had no gross effect on the transcription. Studies on growth of the cells and recovery from ultraviolet damage indicate that the heterologous SSB proteins support DNA replication and recombinational DNA repair of E. coli with the same efficiency as the E. coli SSB protein. Interactions with other E. coli proteins involved in these processes either do not occur, or are not impeded.
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PMID:The single-stranded-DNA-binding proteins (SSB) of Proteus mirabilis and Serratia marcescens. 792 78

Escherichia coli strains carrying mutations in sbcB (exonuclease I) or xthA (exonuclease III) accumulate high-molecular-weight linear plasmid concatemers when transformed with plasmids containing the chi sequence, 5'-GCTGGTGG-3'. Chi-dependent formation of high-molecular-weight plasmid DNA is dependent on recA and recF functions. In addition, chi stimulation occurs only in cis. Our data are consistent with models in which RecA and RecF proteins bind to and protect the DNA ends produced by RecBCD-chi interaction.
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PMID:Chi-dependent formation of linear plasmid DNA in exonuclease-deficient recBCD+ strains of Escherichia coli. 805 Oct 22

We have measured the rates and efficiencies of DNA unwinding (the number of ATP molecules hydrolyzed per DNA base pair unwound) catalyzed by the RecBC,RecBCD-K177Q (a site-directed mutant in the putative ATP-binding site in the RecD subunit), and RecBCD enzymes from Escherichia coli. The DNA unwinding rate was measured with a coupled assay in which unwound DNA is degraded by the combined action of the RecJ enzyme and exonuclease I. The rates of DNA unwinding by the RecBC and RecBCD-K177Q enzymes are reduced by about 4-fold compared to the case of the RecBCD enzyme. The efficiency of ATP hydrolysis was determined in two ways. First, it was calculated from the ratio of the ATP hydrolysis rate to the rate of DNA unwinding. In the second method, ATP hydrolysis was measured under conditions where all of the DNA substrate becomes completely unwound. The efficiency is the ratio of the total amount of ATP hydrolyzed to the amount of DNA substrate present in the reaction. The average efficiencies measured kinetically and by the complete unwinding experiment are as follows: 2.30 and 1.74 ATP/base pair (RecBCD enzyme); 1.44 and 1.28 (RecBC); and 1.20 and 1.07 (RecBCD-K177Q). The RecBC and RecBCD-K177Q enzymes are therefore able to couple ATP hydrolysis to DNA unwinding at least as efficiently as the RecBCD holoenzyme. The lower ATP per base pair ratios found for RecBC and RecBCD-K177Q indicate that the RecD subunit hydrolyzes ATP during DNA unwinding by the RecBCD enzyme.
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PMID:Efficiency of ATP hydrolysis and DNA unwinding by the RecBC enzyme from Escherichia coli. 806 30

The E. coli single-stranded binding protein (SSB) has been demonstrated in vitro to be involved in a number of replicative, DNA renaturation, and protective functions. It was shown previously that SSB can interact with exonuclease I to stimulate the hydrolysis of single-stranded DNA. We demonstrate here that E. coli SSB can also enhance the DNA deoxyribophosphodiesterase (dRpase) activity of exonuclease I by stimulating the release of 2-deoxyribose-5-phosphate from a DNA substrate containing AP endonuclease-incised AP sites, and the release of 4-hydroxy-2-pentenal-5-phosphate from a DNA substrate containing AP lyase-incised AP sites. E. coli SSB and exonuclease I form a protein complex as demonstrated by Superose 12 gel filtration chromatography. These results suggest that SSB may have an important role in the DNA base excision repair pathway.
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PMID:Escherichia coli single-stranded DNA binding protein stimulates the DNA deoxyribophosphodiesterase activity of exonuclease I. 812 10

The combined action of exonuclease I and recA protein leads to a kind of reverse DNA strand exchange in which joint molecules formed on the "wrong" or distal end of a linear duplex in the presence of ATP are stabilized by exonuclease I degradation of the displaced (+) strand. Continued pairing and degradation of the displaced strand leads to strand exchange that appears to progress with a polarity opposite that of the normal recA protein promoted reaction (i.e. 3'-5' with respect to the (+) strand). However, in contrast to the normal 5'-3' strand exchange, the displaced strand is completely degraded in the process. When the linear duplex DNA substrate has a heterologous region at the 5' (proximal) end, the major product (described in a previous study (Bedale, W. A., Inman, R. B., and Cox, M. M. (1991) J. Biol. Chem. 266, 6499-6510)) is a circular duplex DNA molecule with a double-stranded tail whose length corresponds closely to the heterologous segment of the substrate. The origin of this product is here shown to be the result of the exonuclease activity of exonuclease I (either added exogenously or present as a trace contaminant of recA protein or SSB protein preparations), as opposed to endonucleolytic or mechanical breakage. The levels of exonuclease I required to generate these products are sufficiently low that they are undetected by assays for exonuclease contamination in recA protein preparations. These results demonstrate that the interplay of recA protein with other enzymes can have a profound effect on both the mechanism and outcome of recA protein-promoted DNA strand exchange. They also demonstrate that the (+) strand of the duplex DNA substrate is at least transiently displaced in recA protein-mediated pairing even when joint molecules are limited to the distal end.
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PMID:A reverse DNA strand exchange mediated by recA protein and exonuclease I. The generation of apparent DNA strand breaks by recA protein is explained. 832 76

Exonuclease I of E. coli is a 3'-->5' exonuclease acting on single-stranded DNA. We further demonstrate that the enzyme can remove phosphoglycolate groups at 3' termini in DNA. These types of lesions are introduced into DNA by agents that cause oxidative damage such as ionizing radiation. An oligonucleotide substrate pd(T)20[32P]dA was treated with acid to remove the adenine base to generate 3' termini containing 2-deoxyribose-5-phosphate end groups. This substrate was then treated with periodate to generate 3'-phosphoglycoaldehyde groups and was further oxidized with I2 to generate 3'-phosphoglycolate groups. The pd(T)20[32P]PGA substrate was annealed to pd(A)40-60 to produce a double-stranded substrate. Exonuclease I was effective in the removal of the 3'-phosphoglycolate groups from this substrate as determined by HPLC separation. With exonuclease III and endonuclease IV of E. coli, exonuclease I is the third activity found in E. coli that is able to excise deoxyribose-phosphate fragments at 3' termini in DNA. These sugar fragments are blocks to DNA polymerase, and their removal is necessary to complete the base excision repair process.
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PMID:Exonuclease I of Escherichia coli removes phosphoglycolate 3'-end groups from DNA. 836 94

Methyl-directed mismatch repair is initiated by the mismatch-provoked, MutHLS-dependent cleavage of the unmodified strand at a hemimethylated d(GATC) sequence. This reaction is independent of the polarity of the unmodified strand and can occur either 3' or 5' to the mismatch on the unmethylated strand (Au, K. G., Welsh, K., and Modrich, P. (1992) J. Biol. Chem. 267, 12142-12148). The overall repair reaction also occurs without regard to polarity of the unmethylated strand. Both hemimethylated configurations of a linear heteroduplex containing a single d(GATC) sequence are subject to methyl-directed correction in Escherichia coli extracts and in a purified repair system. Repair of both heteroduplex orientations requires MutH, MutL, MutS, DNA helicase II, SSB, and DNA polymerase III holoenzyme, but the two substrates differ with respect to exonuclease requirements for correction. When the unmethylated d(GATC) sequence that directs repair is located 5' to the mismatch on the unmodified strand, mismatch correction requires the 5'--> 3' hydrolytic activity of exonuclease VII or RecJ exonuclease. Repair directed by an unmodified d(GATC) sequence situated 3' to the mismatch depends on the 3'--> 5' activity of exonuclease I. Specific requirements for these activities are evident with circular heteroduplexes containing a single asymmetrically placed d(GATC) sequence, with the requirement for a 5'--> 3' or 3'--> 5' hydrolytic activity being determined by the orientation of the unmethylated strand along the shorter path joining the two sites in the DNA circle. This observation suggests that the methyl-directed repair system utilizes the proximal d(GATC) sequence to direct correction. To our knowledge, these experiments represent the first instance in which exonuclease I, exonuclease VII, and RecJ have been implicated in a particular DNA metabolic pathway.
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PMID:Methyl-directed mismatch repair is bidirectional. 838 65

Using electron microscopy and indirect end-labeling methods, we have examined excision tracts produced by the Escherichia coli methyl-directed mismatch repair system on a closed circular G-T heteroduplex that contains a single d(GATC) site. Despite differing polarities of the unmodified strand in the two hemimethylated derivatives of the heteroduplex, that portion of the unmethylated strand spanning the shorter path between the d(GATC) site and mismatch is targeted for excision in both cases. Mismatch-provoked excision occurring on both hemimethylated DNAs requires DNA helicase II, but exonuclease requirements for the reaction depend on heteroduplex orientation. When the d(GATC) sequence on the unmodified strand resides 3' to the mismatch as viewed along the shorter path, excision requires exonuclease I. Excision occurring on the alternate hemimethylated heteroduplex depends on the 5'--> 3' hydrolytic activity of exonuclease VII. Coupled with the previous demonstration that repair initiates via the mismatch-provoked, MutHLS-dependent incision of the unmethylated strand at a d(GATC) sequence (Au, K.G., Welsh, K., and Modrich, P. (1992) J. Biol. Chem. 267, 12142-12148), these findings indicate an excision mechanism in which helicase II displacement renders the incised strand sensitive to the appropriate single-strand exonuclease. Our data imply that hydrolysis commences at the d(GATC) site, proceeds to a point beyond the mismatch, and terminates at a number of discrete sites within a 100-nucleotide region just beyond this site. The extent of excision is therefore controlled by one or more components of the repair system.
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PMID:Bidirectional excision in methyl-directed mismatch repair. 850 11

In Escherichia coli, chi is a recombination hotspot that stimulates RecBCD-dependent exchange at and to one side of itself. chi activity is highest at chi and decreases with distance from chi. The decrease in chi activity may be a simple property of the physical distance over which chi can stimulate recombination. Alternatively, the decay in chi activity with distance may reflect the high likelihood that chi-stimulated recombination occurs in a single chi-proximal act, to the exclusion of additional chi-stimulated exchanges more distal to chi. To test the models, we determined if chi activity decreases as a function of physical distance (i.e., DNA base pairs) or genetic distance (homologous DNA base pairs). Our results indicate that chi activity decays as a function of genetic distance. In addition, we found that the sbcB gene product (exonuclease I, a 3'-->5' ssDNA exonuclease) modulates the distance over which chi can act. In contrast, the recJ gene product (a 5'-->3' ssDNA exonuclease) does not alter the decay of chi activity.
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PMID:Chi recombination activity in phage lambda decays as a function of genetic distance. 858 27


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