Molecular diversities of RNases H

Ohtani, Naoto; Haruki, Mitsuru; Morikawa, Masaaki; Kanaya, Shigenori

doi:10.1016/s1389-1723(99)80168-6

Cited by 109 publications

(149 citation statements)

References 28 publications

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“…The results indicated that Cpn-RNase HII (CP0654) and CpnRNase HIII (CP0782) both belong to the type 2 RNase H group, which could complement E. coli RNase H mutants in vivo and exhibit RNase H activity in vitro. This is consistent with previous reports that type 2 RNase H could complement RNase HI-deficient E. coli, such as the RNase HII/III from B. subtilis (Ohtani et al, 1999a) or B. stearothermophilus (Chon et al, 2004). Further different properties of the two proteins imply that Cpn-RNase HII may be the major RNase H of C. pneumoniae and the RNase HII/HIII combination in C. pneumoniae AR39 is not a simple substitution of E. coli RNase HI/II.…”

Section: Discussionsupporting

confidence: 93%

“…B. subtilis (Ohtani et al, 1999a) and B. stearothermophilus (Chon et al, 2004). Therefore, the presence of two type 2 RNase Hs in C. pneumoniae AR39 may not be unusual.…”

Section: Discussionmentioning

confidence: 99%

“…Ribonuclease H (RNase H) endonucleolytically hydrolyses the RNA strand of RNA-DNA hybrids in the presence of divalent cations; it is involved in DNA replication, repair, and/or transcription processes and widely present in all three kingdoms (Ohtani et al, 1999a). RNase Hs have been classified into type 1 and type 2 RNase H, and prokaryotic RNase Hs are divided into three groups, RNase HI, HII and HIII, encoded by rnhA, rnhB and rnhC, respectively (Ohtani et al, 1999a, b).…”

Section: Introductionmentioning

confidence: 99%

“…Although many bacteria, such as Escherichia coli, contain both type 1 and type 2 RNase H, some bacteria, such as Bacillus stearothermophilus, Streptococcus pneumoniae, Chlamydia trachomatis and Aquifex aeolicus only have two different type 2 RNase Hs (Ohtani et al, 1999a). Although the crystal structures of some RNase HI and HII enzymes have been determined, e.g.…”

Section: Introductionmentioning

confidence: 99%

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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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Section: Discussionsupporting

confidence: 93%

“…B. subtilis (Ohtani et al, 1999a) and B. stearothermophilus (Chon et al, 2004). Therefore, the presence of two type 2 RNase Hs in C. pneumoniae AR39 may not be unusual.…”

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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

et al. 2007

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“…The enzyme is ubiquitously present in various organisms and is involved in DNA replication and repair. RNase H is classified into two major types according to the sequence similarity, type 1 and type 2 RNase H (22,23). Tk-RNase HII is a member of type 2 RNase H. A comparison of the crystal structures of Escherichia coli RNase HI, Thermus thermophilus RNase HI, and Tk-RNase HII indicates that these enzymes share a folding motif, referred to as the RNase H-fold due to its characterization in E. coli RNase HI (24), although there is no sequence similarity between RNase HI and RNase HII.…”

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confidence: 99%

Kinetically Robust Monomeric Protein from a Hyperthermophile

et al. 2004

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Equilibrium and kinetic studies were carried out under denaturation conditions to clarify the energetic features of the high stability of a monomeric protein, ribonuclease HII, from a hyperthermophile, Thermococcus kodakaraensis (Tk-RNase HII). Guanidine hydrochloride (GdnHCl)-induced unfolding and refolding were measured with circular dichroism at 220 nm, and heat-induced denaturation was studied with differential scanning calorimetry. Both GdnHCl-and heat-induced denaturation are very reversible. It was difficult to obtain the equilibrated unfolding curve of Tk-RNase HII below 40°C, because of the remarkably slow unfolding. The two-state unfolding and refolding reactions attained equilibrium at 50°C after 2 weeks. The Gibbs energy change of GdnHCl-induced unfolding (∆G(H 2 O)) at 50°C was 43.6 kJ mol -1 . The denaturation temperature in the DSC measurement shifted as a function of the scan rate; the denaturation temperature at a scan rate of 90°C h -1 was higher than at a scan rate of 5°C h -1 . The unfolding and refolding kinetics of Tk-RNase HII were approximated as a first-order reaction. The ln k u and ln k r values depended linearly on the denaturant concentration between 10 and 50°C. The ∆G(H 2 O) value obtained from the rate constant in water using the two-state model at 50°C, 44.5 kJ mol -1 , was coincident with that from the equilibrium study, 43.6 kJ mol -1 , suggesting the two-state folding of TkRNase HII. The values for the rate constant in water of the unfolding for Tk-RNase HII were much smaller than those of E. coli RNase HI and Thermus thermophilus RNase HI, which has a denaturation temperature similar to that of Tk-RNase HII. In contrast, little difference was observed in the refolding rates among these proteins. These results indicate that the stabilization mechanism of monomeric protein from a hyperthermophile, Tk-RNase HII, with reversible two-state folding is characterized by remarkably slow unfolding.Microorganisms can be classified into psychrophiles, mesophiles, thermophiles, or hyperthermophiles on the basis of the differences in their optimal growth temperatures. The stabilities of proteins from these microorganisms exhibit different properties. Proteins from hyperthermophiles generally reveal greater stability than those from any other microorganisms. Therefore, proteins from hyperthermophiles are expected to provide unique information about protein folding, stability, and function (1-7).The stability of proteins in solution is evaluated by the Gibbs energy changes (∆G) upon denaturation, when they are reversible under experimental conditions. The value of ∆G is obtained from equilibrium and kinetic experiments. Studies concerning the stability of proteins from hyperthermophiles have focused primarily on the equilibrium aspects (8-13). The results suggested that extremely high stability of these proteins can be achieved by increasing the number of ionic interactions and the extent of hydrophobic surface burial. Several reports have recently appeared in which the equilibrium...

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A Simple Colorimetric Au‐on‐Au Tip Sensor with a New Functional Nucleic Acid Probe for Food‐borne Pathogen Salmonella typhimurium

Khan

et al. 2023

Angewandte Chemie

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An Au‐on‐Au tip sensor is developed for the detection of Salmonella typhimurium (Salmonella), using a new synthetic nucleic acid probe (NAP) as a linker for the immobilization of a DNA‐conjugated Au nanoparticle (AuNP) onto a DNA‐attached thin Au layer inside a pipette tip. In the presence of Salmonella, RNase H2 from Salmonella (STH2) cleaves the NAP and the freed DNA‐conjugated AuNP can be visually detected by a paper strip. This portable biosensor does not require any electronic, electrochemical or optical equipment. It delivers a detection limit of 3.2×103 CFU mL−1 for Salmonella in 1 h without cell‐culturing or signal amplification and does not show cross‐reactivity with several control bacteria. Further, the sensor reliably detects Salmonella spiked in food samples, such as ground beef and chicken, milk, and eggs. The sensor can be reused and is stable at ambient temperature, showing its potential as a point‐of‐need device for the prevention of food poisoning by Salmonella.

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Molecular diversities of RNases H

Cited by 109 publications

References 28 publications

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

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

Kinetically Robust Monomeric Protein from a Hyperthermophile

A Simple Colorimetric Au‐on‐Au Tip Sensor with a New Functional Nucleic Acid Probe for Food‐borne Pathogen Salmonella typhimurium

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