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PubMed1999

Molecular modeling of the three-dimensional architecture of the RNA component of yeast RNase MRP.

RNase mitochondrial RNA processing (MRP) is a ribonucleoprotein endoribonuclease that is involved in RNA processing events in both the nucleus and the mitochondria. The MRP RNA is both structurally and evolutionarily related to RNase P, the ribonucleoprotein endoribonuclease that processes the 5'-end of tRNAs. Previous analysis of the RNase MRP RNA by phylogenetic analysis and chemical modification has revealed strikingly conserved secondary structural elements in all characterized RNase MRP RNAs. Utilizing successive constraint modeling and energy minimization I derived a three-dimensional model of the yeast RNase MRP RNA. The final model predicts several notable features. First, the enzyme appears to contain two separate structural domains, one that is highly conserved among all MRP and P RNAs and a second that is only conserved in MRP RNAs. Second, nearly all of the highly conserved nucleotides cluster in the first domain around a long-range interaction (LRI-I). This LRI-I is characterized by a ubiquitous uridine base, which points into a cleft between these two structural domains generating a potential active site for RNA cleavage. Third, helices III and IV (the yeast equivalent of the To-binding site) model as a long extended helix. This region is believed to be the binding site of shared proteins between RNase P and RNase MRP and would provide a necessary platform for binding these seven proteins. Indeed, several residues conserved between the yeast MRP and P RNAs cluster in the central region of these helixes. Lastly, characterized mutations in the MRP RNA localize in the model based on their severity. Those mutations with little or no effect on the activity of the enzyme localize to the periphery of the model, while the most severe mutations localize to the central portion of the molecule where they would be predicted to cause large structural defects. Press.

Bacillus subtilis↗
PubMed1999

Similarities and differences in the RNase H activities of human immunodeficiency virus type 1 reverse transcriptase and Moloney murine leukemia virus reverse transcriptase.

Retroviral revXerse transcriptases (RTs) have an associated RNase H activity that can cleave RNA-DNA duplexes with considerable precision. We believe that the structure of the RNA-DNA duplexes in the context of RT determines the specificity of RNase H cleavage. To test this idea, we treated three related groups of synthetic RNA-DNA hybrids with either Moloney murine leukemia virus (MLV) RT or human immunodeficiency virus type 1 (HIV-1) RT. All of the hybrids were prepared using the same 81-base RNA template. The first series of RNase H substrates was prepared with complementary DNA oligonucleotides of different lengths, ranging from 6 to 20 nucleotides, all of which shared a common 5' end and were successively shorter at their 3' ends. The second series of oligonucleotides had a common 3' end but shorter 5' ends. The DNA oligonucleotides in the third series were all 20 bases long but had non-complementary stretches at either the 5' end, 3' end, or both ends. Several themes have emerged from the experiments with these RNA-DNA duplexes. (1) Both HIV-1 RT and MLV RT cleave fairly efficiently if the duplex region is at least eight bases long, but not if it is shorter. (2) Although, under the conditions we have used, both enzymes require the substrate to have a region of RNA-DNA duplex, both MLV RT and HIV-1 RT can cleave RNA outside the region that is part of the RNA-DNA duplex. (3) The polymerase domain of HIV-1 RT uses certain mismatched segments of RNA-DNA to position the enzyme for RNase H cleavage, whereas the polymerase domain of MLV RT does not use the same mismatched segments to define the position for RNase H cleavage. (4) For HIV-1 RT, a mismatched region near the RNase H domain can interfere with RNase H cleavage; cleavage is usually (but not always) more efficient if the mismatched segment is deleted. These results are discussed in regard to the structure of HIV-1 RT and the differences between HIV-1 RT and MLV RT.

Amino Acid Sequence↗
PubMed2002

tRNA 3' end maturation in archaea has eukaryotic features: the RNase Z from Haloferax volcanii.

Here, we report the first characterization and partial purification of an archaeal tRNA 3' processing activity, the RNase Z from Haloferax volcanii. The activity identified here is an endonuclease, which cleaves tRNA precursors 3' to the discriminator. Thus tRNA 3' processing in archaea resembles the eukaryotic 3' processing pathway. The archaeal RNase Z has a KCl optimum at 5mM, which is in contrast to the intracellular KCl concentration being as high as 4M KCl. The archaeal RNase Z does process 5' extended and intron-containing pretRNAs but with a much lower efficiency than 5' matured, intronless pretRNAs. At least in vitro there is thus no defined order for 5' and 3' processing and splicing. A heterologous precursor tRNA is cleaved efficiently by the archaeal RNase Z. Experiments with precursors containing mutated tRNAs revealed that removal of the anticodon arm reduces cleavage efficiency only slightly, while removal of D and T arm reduces processing effciency drastically, even down to complete inhibition. Comparison with its nuclear and mitochondrial homologs revealed that the substrate specificity of the archaeal RNase Z is narrower than that of the nuclear RNase Z but broader than that of the mitochondrial RNase Z.

Anticodon↗
PubMed1990

Captan binding to avian myeloblastosis virus reverse transcriptase and its effect on RNase H activity.

The inhibitor captan (N-trichloromethylthio-4-cyclohexen-1,2-dicarboximide) was used to explore the ribonuclease H (RNase H) active site of avian myeloblastosis virus (AMV) reverse transcriptase. Gel permeation chromatography of purified enzyme showed that [14C]captan bound to the alpha subunit in a ratio of 10:1 and to a 32,000 d polypeptide in a ratio of 4:1. Neither the alpha beta nor the beta subunit bound [14C]captan. The binding of 5 of the captan molecules was prevented by preincubating enzyme with polynucleotide. Deoxyguanosine triphosphate (dGTP) protected the enzyme against the binding of 4 captan molecules. Each holoenzyme bound 2 molecules of [3H]dGTP in the absence of, and 1 molecule of [3H]dGTP in the presence of 1 mM captan. Ribonuclease H activity was inhibited when AMV reverse transcriptase was preincubated with 1 mM captan before the degradative reaction was initiated. Preincubation of enzyme with polynucleotide before exposure to captan could partially protect the RNase H activity (61 +/- 2% activity remained). Deoxyguanosine triphosphate also partially protected the RNase H activity from inhibition by captan (75 +/- 9% activity remained). Inhibition of the RNase H activity was completely prevented by preincubating enzyme simultaneously with polynucleotide and dGTP. When separated by glycerol gradients the alpha subunit and alpha beta dimer both exhibited RNase H activity, but only the RNase H activity of the alpha subunit was inhibited by captan. Activity and binding studies revealed that the RNase H and polymerase activities of the alpha subunit are not susceptible to the interaction of captan when this subunit is in the alpha beta dimer form.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMedDate not supplied

RNase P from bacteria. Substrate recognition and function of the protein subunit.

RNase P recognizes many different precursor tRNAs as well as other substrates and cleaves all of them accurately at the expected position. RNase P recognizes the tRNA structure of the precursor tRNA by a set of interactions between the catalytic RNA subunit and the T- and acceptor-stems mainly, although residues in the 5'-leader sequence as well as the 3'-terminal CCA are important. These conclusions have been reached by several studies on mutant precursor tRNAs as well as cross-linking studies between RNase P RNA and precursor tRNAs. The protein subunit of RNase P seems also to affect the way that the substrate is recognized as well as the range of substrates that can be used by RNase P, although the protein does not seem to interact directly with the substrates. The interaction between the protein and RNA subunits of RNase P has been extensively studied in vitro. The protein subunit sequence is not highly conserved among bacteria, however different proteins are functionally equivalent as heterologous reconstitution of the RNase P holoenzyme can be achieved in many cases.

Amino Acid Sequence↗
PubMed2004

Isolation, characterization, and evolutionary divergence of mouse RNase 6: evidence for unusual evolution in rodents.

The evolution of the ribonuclease A (RNase A) vertebrate-specific enzyme family is interesting in that specific gene lineages appear to be responding to unique selective pressures in wildly diverse manners to generate proteins that are capable of reducing the infectivity of viruses, killing systemic pathogens, and inducing the growth of blood vessels all while maintaining the signature motifs of a ribonuclease. In this paper, we present the DNA sequence and gene structure of Mus musculus RNase 6 and examine the expression pattern and enzymatic activity of the recombinant protein. M. musculus RNase 6 has a limited expression pattern compared to human RNase 6 and is an efficient ribonuclease, with a catalytic efficiency 17-fold higher than that of human protein. Evolutionary analysis reveals that RNase 6 was subject to unusual evolutionary forces (dN/dS = 1.2) in an ancestral rodent lineage before the separation of Mus and Rattus. However, more recent evolution of rodent RNase 6 has been relatively conserved, with an average dN/dS of 0.66. These data suggest that the ancestral rodent RNase 6 was subject to accelerated evolution, resulting in the conserved modern gene, which most likely plays an important role in mouse physiology.

Amino Acid Sequence↗
PubMed2002

Evolutionary analysis of S-RNase genes from Rosaceae species.

Eight new cDNA sequences for S-RNases were cloned and analysed from almond (Prunus dulcis) cultivars of European origin, and compared to published sequences from other Rosaceae species. Insertions/deletions of 10-20 amino acid residues were detected in the RC4 and C5 domains of S-RNases from almond and sweet cherry. The S-RNases of the Prunus species and those of the genera Malus and Pyrus formed two distinct groups on phylogenetic analysis. Nucleotide substitutions were analysed in the S-RNase genes of these species. The S-genes of almond and sweet cherry have a lower Ka/Ks value than those of apple, pear and wild apple do. The fact that there is no fixed difference between the S-RNase genes of almond and sweet cherry, or between apple and pear, suggests that nucleotide substitutions only introduce transient polymorphism into the two groups, and rarely became fixed and contribute to divergence. Through the comparative study of 17 S-RNase genes from the genus Prunus and 18 from the genera Malus and Pyrus, some fixed nucleotide differences between the two groups were identified. These differences do not appear to be the result of selection for adaptive mutations, since the number of replacement substitutions is not significantly greater than the number of synonymous substitutions. S-RNase genes of almond and sweet cherry, and of apple and pear, showed little heterogeneity in nucleotide substitution rates. However, heterogeneity was observed between the two groups of S-alleles, with the Prunus alleles exhibiting a lower rate of non-synonymous substitutions than alleles from Malus and Pyrus. The evolutionary relationships between these species are discussed.

Amino Acid Sequence↗
PubMed2006

Organization of the interferon-inducible 2',5'-oligoadenylate-dependent ribonuclease L (RNase L) gene of mouse.

Interferons (IFNs) induce a 2',5'-oligoadenylate (2-5A)-dependent ribonuclease L (RNase L) following virus-infection of mammalian cells. RNase L degrades both viral and cellular RNAs and restricts virus-proliferation. We have studied organization of RNase L gene in genomic DNA from the mouse liver by Southern blot analysis. Several BamHI, BglII, EcoRI, HincII, HindIII, NcoI, PstI, SacI, and XbaI restriction fragments hybridized to (32)P-labeled mouse RNase L cDNA and the 5'-proximal exon probes. Mouse RNase L gene exists as a single copy (>16 kb DNA) gene. A 5 kb HindIII and a 2.5 kb EcoRI DNA were detected as 5'-upstream DNA of the gene which may contain mouse RNase L promoter. Our results will help studying mouse RNase L gene promoter in further details.

5' Flanking Region↗
PubMed1990

Photochemical crosslinking in oligonucleotide-protein complexes between a bromine-substituted 2-5A analog and 2-5A-dependent RNase by ultraviolet lamp or laser.

2',5'-oligoadenylates known as 2-5A [px(A2'p)nA; chi = 2 or 3, n greater than or equal to 2] are produced in interferon-treated cells in response to double-stranded RNA. 2-5A binds with high affinity to a 2-5A-dependent RNase resulting in the cleavage of single-stranded RNA. An efficient, rapid, and extremely sensitive photoaffinity labeling method was developed to facilitate detection of 2-5A-dependent RNase. A bromine-substituted and radioactive derivative of 2-5A, the 5'-monophosphate, p(A2'p)2(br8A2'p)2A3'-[32P]Cp, was synthesized as probe for 2-5A-dependent RNase. Even though this bromine-substituted analog of 2-5A bore no 5'-terminal triphosphate or diphosphate, it bound to 2-5A-dependent RNase with the same high affinity as did 2-5A per se but it was a less effective activator of the RNase under the present assay conditions. The presence of bromine atoms in the 2-5A analog enhanced by more than 200-fold crosslinking to 2-5A-dependent RNase under a uv lamp; many additional polypeptides were also labeled but at much lower levels. Furthermore, using high-intensity uv laser irradiation (308 nm) covalent attachment of the bromine-substituted 2-5A analog to 2-5A-dependent RNase was readily achieved within 10(-6) s.

Adenine Nucleotides↗
PubMed1991

Regeneration of enzyme activity after western blot: activation of RNase L by 2-5A on filter--importance for its detection.

A rapid and convenient new procedure for detecting RNase L activity following Western blot by renaturation of the enzyme on the nitrocellulose sheets is described. This method allows the simultaneous analysis of enzymatic activity (e.g., cleavage of poly(uridylic acid)-3'-[32P]pCp) and RNase L binding to radioactivE probes (e.g., 2-5A-3'-[32P]pCp) in the same sample. Unlike previously published methods, this procedure eliminates interference from proteases or other RNases during the analysis of RNase L activity. The detection of RNase(s) L is also affected by the presence of endogenous 2-5A, 2-5A derivatives, or other possible "inhibitors" in cell extracts; this Western blot assay allows of RNase(s) L to be detected independently of intracellular 2-5A or analogs. Differences between the procedures used so far and this Western blot technique can indeed be demonstrated. It is shown with this Western blot assay that although RNase L has been described as a protein of 185-200 kDa under nondenaturating conditions, its 80-kDa (and 40-kDa) component is able to bind 2-5A and to cleave poly(uridylic acid) in a 2-5A-dependent way, independently of other subunit(s) or cofactor(s).

Adenine Nucleotides↗
PubMed1992

Affinity blotting assay for 2-5A-dependent RNase.

The intracellular effectors known as 2-5A (ppp-(A2'p)nA) regulate the cleavage of single-stranded RNA by activating a latent endoribonuclease (2-5A-dependent RNase or RNase L). Accordingly this enzyme may exist in either an inactive form, free of 2-5A, or an active, 2-5A-bound form. Previously, a radiobinding assay for 2-5A-dependent RNase was developed that measured the amount of labeled ppp(A2'p)2A3'-[32P]Cp, a derivative of ppp(A2'p)nA, that bound the inactive enzyme form. Because 2-5A-dependent RNase has a particularly high affinity for 2-5A the radiobinding assay may not measure the 2-5A activated form of the enzyme. Therefore an efficient procedure to facilitate the detection of total 2-5A-dependent RNase (i.e., 2-5A-free and 2-5A-bound enzyme) in mouse spleen extracts was developed. Denaturing conditions were used to ensure that all 2-5A-dependent RNase was in the 2-5A-free form. After denaturation on polyacrylamide gel electrophoresis, optimal blotting conditions onto nitrocellulose and renaturation of the 2-5A binding site of 2-5A-dependent RNase were developed. This procedure allowed a population of enzyme that otherwise is not accessible by the classical radiobinding assay to be assayed, thus leading to an increased measurement of 15-17% in cytoplasmic spleen extracts.

Affinity Labels↗
PubMed1992

Effects of glycosylation on protein conformation and amide proton exchange rates in RNase B.

Assignment of most of the proton NMR resonances of bovine pancreatic RNase B has been achieved using standard NMR techniques and by comparison with the published assignments for RNase A. A comparison of the NMR spectra of RNase B with RNase A shows that glycosylation of the enzyme has little overall effect on the conformation of the protein in solution. Comparisons of hydrogen-deuterium solvent exchange rates for the NH protons of RNase A and RNase B were made using two-dimensional 1H correlation spectroscopy. In the case of the glycosylated enzyme the exchange rates decreased for the NH protons of residues 9-14, 23-24, 32, 34-35, 39-40, 43-44, 48-49, 60, 71, 75-76, 80, 83-85, 100-101, 107, 111 and 122, relative to the unglycosylated RNase A. These results are consistent with the presence of the oligosaccharide inducing enhanced global dynamic stability and consequent changes to the unfolding equilibrium of the enzyme. The enhanced stability is observed not only for residues in the vicinity of the glycosylation site, asparagine-34, but also at residues remote from this site, as much as 30 A away.

Amides↗
PubMed1987

Analysis of RNase-A-resistant regions of adenovirus 2 major late precursor-mRNA in splicing extracts reveals an ordered interaction of nuclear components with the substrate RNA.

An RNase A protection assay was employed to investigate the interaction of nuclear components with a precursor-mRNA derived from the adenovirus 2 major late transcription unit in a splicing extract from HeLa cells. Upon incubation in the extract, two regions in the precursor-RNA become resistant to digestion with RNase A. After short incubation times (5 min) at 30 degrees C, fragments mapping upstream from the branch point in the intron are obtained. After ten minutes or more, additional oligonucleotides, derived from the 5' splice site, are protected. RNase A protection of different RNA substrates demonstrates that a 5' splice site is not required for the binding of components to the branch point region. For interaction with this site, the polypyrimidine stretch just upstream from the 3' splice site is essential. Binding to the 5' splice site occurs only in the presence of an intact 3' end of the intron. Preincubation of the extract with excess unlabelled RNA containing only a 3' splice site leads to efficient competition of binding, both in the branch point region and at the 5' splice site, whereas an RNA that contains only 5'-splice-site sequences has no effect on the interaction with the mRNA precursor. This indicates that stable association with the 5' splice site requires prior binding of components in the branch point region. When splicing complexes are digested with RNase A, it becomes apparent that only the branch point region is sequestered into a ribonucleoprotein (RNP) structure in the 35 S complex. The 5' splice site becomes resistant to RNase A only when a 50 S splicing complex has been assembled. Degradation of specific regions in U1, U2 and U4 RNA with complementary oligodeoxynucleotides and RNase H has been used to analyse involvement of the U small nuclear RNPs (snRNPs) in the protection reaction. The 5' end of U2 RNA is essential for protection of the branch point region. RNA sequences in a loop of U2 RNA (nucleotides 65 to 78) are required for the formation of an RNase-A-resistant structure at the 5' splice site. Taken together, these results suggest that U2 snRNP participates in the formation of a pre-splicing complex, the 5' end of its RNA being involved in the observed binding. Conversion to a 50 S splicing complex is obtained after the binding of U1 and U4/U6 snRNPs, which also requires sequences in a loop of U2 RNA. Possible interactions between the individual snRNPs and between snRNPs and precursor-mRNA are discussed.

Adenoviridae↗
PubMed1996

Epidermal growth factor receptor-dependent cytotoxicity for human squamous carcinoma cell lines of a conjugate composed of human EGF and RNase 1.

Recombinant human ribonuclease 1 (RNase 1) was chemically linked to recombinant human epidermal growth factor (EGF). The EGF-RNase conjugate showed dose-dependent cytotoxicity for EGF receptor-overexpressing A431 and TE-8 human squamous carcinoma cells with an IC50 of 2 x 10(-7)M and 10(-6)M, respectively, whereas the IC50 of RNase alone was almost 10(-4)M. An unconjugated mixture of EGF and RNase had no greater effect than RNase alone. The conjugate showed no detectable cytotoxicity against EGF receptor-deficient small cell lung cancer cells (H69). Addition of excess EGF in the medium protected A431 cells from the EGF-RNase conjugate cytotoxicity. The cytotoxicity of the EGF-RNase conjugate was positively correlated with the EGF receptor numbers of each cell line. The chimeric toxin composed of only human proteins might be a more useful anti-cancer agent with less immunogenicity than the conventional chimeric toxins.

Carcinoma, Squamous Cell↗
PubMed1993

NMR studies of a complex of RNAse from Penicillium brevicompactum with dinucleoside phosphonate and the implications for the mechanism of enzyme action.

The chemical-shift dependences of the proton signals of the guanosine and uridine moieties were measured as a function of the relative amount of GpcU complexed with RNase Pb1 (EC 3.1.27.3). The equal values of the chemical-shift changes of the guanosine C8-protons on complex formation between GpcU and RNase Pb1 and that of the 3'-GMP and RNase Pb1 allow to conclude that the guanosine base is bound in the same manner in these protein-ligand complexes. The guanosine moiety of GpcU is also most probably bound in the syn-conformation. The absence of changes in both the linewidths and the chemical shifts of the C1', C5 and C6-proton signals of the uridine on complex formation indicates that the uridine moiety of the dinucleoside phosphonate is not immobilized in the complex. The pH dependences of the chemical shifts of the C2-protons of the histidine-imidazole ring of RNase Pb1 and that of the 31P of GpcU in the RNase complex were studied. The results suggest that there is a direct interaction between the phosphonate group of the ligand and the protonated imidazole ring of His-90. The side groups of His-38 and Glu-56 are hydrogen bonded to each other at neutral pH and they are located in the vicinity of the phosphonate group of GpcU. When the carboxyl group of Glu-56 is protonated the His-38 imidazole ring forms a new hydrogen bond with one of the phosphoryl oxygens of the phosphonate group. On the basis of these results we propose the mechanism of action of RNase Pb1 which is probably also true for RNase T1.

Aspergillus↗
PubMed1996

Expression of the mitochondrial RNase P RNA subunit-encoding gene from a variant promoter sequence in Saccharomyces cerevisiae.

Ribonuclease P (RNase P) is a common tRNA processing enzyme that removes the 5' leader sequence of precursor tRNAs. This activity is identified in yeast mitochondria as a separate enzyme from the nuclear RNase P. Like other RNase P enzymes, the mitochondrial (mt) RNase P is also a ribonucleoprotein composed of both RNA and protein subunits. The RNA subunit is encoded by a mt gene and the protein subunit is supplied by a nuclear gene. Earlier studies described one active promoter (FP1) located 5' to the mt tRNA(fMet)-RNase P RNA-tRNA(Pro) gene cluster, so that the mitochondrially encoded RNA subunit was thought to be co-transcribed with two of its substrate tRNAs. However, the results of in vitro transcription and primer extension experiments presented here demonstrate that the mt RNase P RNA subunit-encoding gene (RPM1) is transcribed from a new promoter (SP)which is located between the tRNA(fMet) and RPM1 genes. The sequence [5'-TATAAGAA(+1)] of the new promoter varies from the conserved promoter sequence [5'-TATAAGTA(+1)], but is one of the sequences that is active in the in vitro transcription assay to determine the consensus promoter sequence [5'-T A T/a A A/g/c G T/a/c N(+1)]. This result demonstrates that a naturally occurring variant promoter is used by RPM1. Identification of the novel SP promoter suggests that the synthesis of the mt RNase P RNA subunit might be uncoupled from the expression of upstream tRNA(fMet) gene, and that RPM1 might be independently transcribed in Saccharomyces cerevisiae.

Animals↗
PubMed2006

Solution structure of DNA/RNA hybrid duplex with C8-propynyl 2'-deoxyadenosine modifications: Implication of RNase H and DNA/RNA duplex interaction.

Solution structures of DNA/RNA hybrid duplexes, d(GCGCA*AA*ACGCG): r(cgcguuuugcg)d(C) (designated PP57), containing two C8-propynyl 2'-deoxyadenosines (A*) and unmodified hybrid (designated U4A4) are solved. The C8-propynyl groups on 2'-deoxyadenosine perturb the local structure of the hybrid duplex, but overall the structure is similar to that of canonical DNA/RNA hybrid duplex except that Hoogsteen hydrogen bondings between A* and U result in lower thermal stability. RNase H is known to cleave RNA only in DNA/RNA hybrid duplexes. Minor groove widths of hybrid duplexes, sugar puckerings of DNA are reported to be responsible for RNase H mediated cleavage, but structural requirements for RNase H mediated cleavage still remain elusive. Despite the presence of bulky propynyl groups of PP57 in the minor groove and greater flexibility, the PP57 is an RNase H substrate. To provide an insight on the interactions between RNase H and substrates we have modeled Bacillus halodurans RNase H-PP57 complex, our NMR structure and modeling study suggest that the residue Gly(15) and Asn(16) of the loop residues between first beta sheet and second beta sheet of RNase HI of Escherichia coli might participate in substrate binding.

DNA↗
PubMed2004

Activity of the isolated HIV RNase H domain and specific inhibition by N-hydroxyimides.

This report describes a procedure to generate enzymatically active, isolated HIV RNase H domain. In contrast to previously described preparations, the RNA cleavage activity of the untagged RNase H domain was surprisingly similar to that of the full-length HIV-RT protein. Signature cleavages at 18 and 9 nucleotides downstream of a recessed RNA 5'-end were retained with the isolated RNase H domain. Activity was strongly decreased by deletion of 3 amino acids from the C-terminus, consistent with an important structural or functional role of the C-terminal alpha-helix. A prototype N-hydroxyimide (2-hydroxy-4H-isoquinoline-1,3-dione) was found to inhibit the activity of the isolated HIV RNase H domain as well as the RNase H activity of full-length HIV reverse transcriptase. In contrast, the compound did not significantly inhibit the structurally closely related Escherichia coli RNase HI. Specific binding of N-hydroxyimide compounds to the isolated RNase H domain was observed by protein fluorescence quenching.

Amino Acid Substitution↗