Three-dimensional structure of ribonuclease H from E. coli

Katayanagi, Katsuo; Miyagawa, M.; Matsushima, Masaaki; Ishikawa, Mariko; Kanaya, Shigenori; Ikehara, Morio; Matsuzaki, Toshiake; Morikawa, Kosuke

doi:10.1038/347306a0

Cited by 353 publications

(302 citation statements)

References 22 publications

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“…Although the primary amino acid sequence did not provide any clue to the function of PF2046, analysis of the structure of PF2046 by DALI, Profunc and CATH indicated that the secondary structural elements of PF2046 are similar to a RNase HI (Katayanagi et al, 1990;Davies et al, 1991;Ohtani et al, 2004). We analyzed the structure further for signature motifs and catalytic residues necessary for the processing of nucleic acids.…”

Section: Discussionmentioning

confidence: 99%

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Crystal structure of a novel non-Pfam protein PF2046 solved using low resolution B-factor sharpening and multi-crystal averaging methods

Shaw

et al. 2010

Protein Cell

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Sometimes crystals cannot diffract X-rays beyond 3.0 Å resolution due to the intrinsic flexibility associated with the protein. Low resolution diffraction data not only pose a challenge to structure determination, but also hamper interpretation of mechanistic details. Crystals of a 25.6 kDa non-Pfam, hypothetical protein, PF2046, diffracted X-rays to 3.38 Å resolution. A combination of SeMet derived heavy atom positions with multiple cycles of B-factor sharpening, multi-crystal averaging, restrained refinement followed by manual inspection of electron density and model building resulted in a final model with a R value of 23.5 (R free = 24.7). The asymmetric unit was large and consisted of six molecules arranged as a homodimer of trimers. Analysis of the structure revealed the presence of a RNA binding domain suggesting a role for PF2046 in the processing of nucleic acids.

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Section: Discussionmentioning

confidence: 99%

“…3A). The carboxylic acids are known to bind metal ions in RNase HI (Katayanagi et al, 1990;Davies et al, 1991;Ohtani et al, 2004). In addition to the carboxylic acids, a histidine residue participates in the catalysis.…”

Section: Discussionmentioning

confidence: 99%

Crystal structure of a novel non-Pfam protein PF2046 solved using low resolution B-factor sharpening and multi-crystal averaging methods

Shaw

et al. 2010

Protein Cell

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“…A 12 bp RNA-DNA (59-32 P-cggagaugacgg-39, 39-GCCTCTACTGCC-59; note that ribonucleotides are shown by lower-case letters), 35 bp DNA-RNA-DNA/DNA (59-32 P-AGAGGCAGGAGAGTuGCAACGCAGCAAGAGGACGC-39, 39-TCTCCG-TCCTCTCAACGTTGCGTCGTTCTCCTGCG-59) and Ribo primer (59-32 P-GCGTCCTCTT GCTGCGTTGCaa-39, 39-CGCAGGAGAAC-GACGCAACGTTGAGAGGACGGAGA-59) were used as substrates. All oligonucleotides carrying ribonucleotide(s) were 59-end-labelled with 32 P and the hybrid duplexes were prepared by standard procedures (Katayanagi et al, 1990). The cleavage assays were performed in reaction mixture containing 0.2 mM of different ribonucleotide substrates and specified amounts of Cpn-RNase Hs.…”

Section: In Vitro Enzymic Activity Assaysmentioning

confidence: 99%

“…Although the crystal structures of some RNase HI and HII enzymes have been determined, e.g. E. coli (Katayanagi et al, 1990;Yang et al, 1990), Methanococcus jannaschii (Lai et al, 2000) and Thermoccus kadakaraensis (Muroya et al, 2001), little 3D structural information on RNase HIII is available. The enzymic properties of a few RNase HIIIs such as Bacillus subtilis RNase HIII and B. stearothermophilus RNase HIII have been analysed (Ohtani et al, 1999b;Chon et al, 2004Chon et al, , 2006a.…”

Section: Introductionmentioning

confidence: 99%

Biochemical characterization and functional complementation of ribonuclease HII and ribonuclease HIII from Chlamydophila pneumoniae AR39

et al. 2007

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Chlamydophila pneumoniae AR39 contains two different ORFs (CP0654 and CP0782) encoding ribonuclease H (RNase H) homologues, Cpn-RNase HII and Cpn-RNase HIII. Sequence alignments show that the two homologues both contain the conserved motifs of type 2 RNase H, and Cpn-RNase HII has the conserved active-site motif (DEDD) of RNase HII. Cpn-RNase HIII also contains a unique active-site motif (DEDE), common to other RNase HIIIs. Complementation assays indicated that Cpn-RNase HII can complement both Escherichia coli RNase HII and RNase HI, but Cpn-RNase HIII can only complement the latter. In vitro enzyme activity experiments showed that neither Cpn-RNase HII nor Cpn-RNase HIII is thermostable and their optimum pH values were 9.0 and 10.0, respectively. Cpn-RNase HII cleaves a 12 bp RNA-DNA substrate at multiple sites, but Cpn-RNase HIII at only one site. When a 35 bp DNA-RNA-DNA/DNA chimeric substrate was used, cleavage was only observed with Cpn-RNase HII. These results indicate that the RNase H combination of C. pneumoniae AR39 is not simple substitution of E. coli RNase H, perhaps representing a more primordial type. This is believed to be the first in vivo functional study of Chlamydophila RNase Hs and the results should cntribute to the analysis of RNase Hs of other parasite species.

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“…Among these RNases, RNase H (Crouch, 1990;Katayanagi et al, 1990) has been defined as an enzyme that specifically degrades the ribonucleotide moiety in RNA-DNA hybrid molecules (Ohtani et al, 1999a, b). RNases H are classified into two major families, type I and type II, which are evolutionarily unrelated.…”

Section: Introductionmentioning

confidence: 99%

lmo1273, a novel gene involved in Listeria monocytogenes virulence

et al. 2009

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Listeria monocytogenes is a foodborne pathogen able to infect humans and many other mammalian species, leading to serious, often fatal disease. We have previously identified a five-gene locus in the genome of L. monocytogenes EGD-e which comprised three contiguous genes encoding paralogous type I signal peptidases. In the present study, we focused on the two distal genes of the locus (lmo1272 and lmo1273), encoding proteins sharing significant similarities with the YlqF and RnhB proteins, respectively, of Bacillus subtilis. lmo1273 could complement an Escherichia coli rnhA-rnhB thermosensitive growth phenotype, suggesting that it encodes a functional RNase H. Strikingly, inactivation of lmo1273 provoked a strong attenuation of virulence in the mouse model, and kinetic studies in infected mice revealed that multiplication of the lmo1273 mutant in target organs was significantly impaired. However, the mutation did not impair L. monocytogenes intracellular multiplication or cell-to-cell spread in cell culture models. Transcriptional profiles obtained with an lmo1273-overexpressing strain were compared to those of the wild-type strain, using microarray analyses. The data obtained suggest a pleiotropic regulatory role of Lmo1273 and possible links with amino acid uptake.

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Three-dimensional structure of ribonuclease H from E. coli

Cited by 353 publications

References 22 publications

Crystal structure of a novel non-Pfam protein PF2046 solved using low resolution B-factor sharpening and multi-crystal averaging methods

Crystal structure of a novel non-Pfam protein PF2046 solved using low resolution B-factor sharpening and multi-crystal averaging methods

Biochemical characterization and functional complementation of ribonuclease HII and ribonuclease HIII from Chlamydophila pneumoniae AR39

lmo1273, a novel gene involved in Listeria monocytogenes virulence

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