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: UMLS:C0679427 (myeloblastosis)
982 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Deoxyribonucleotide insertion efficiencies were measured opposite site-directed abasic template lesions using human immunodeficiency virus 1 reverse transcriptase (HIV-1RT), and the efficiencies to continue primer synthesis beyond the lesion, by addition of the "next correct" deoxynucleotide, were measured as a function of sequence context. Insertion of purines was favored over pyrimidines, A > G > T approximately C. Primer extension past the lesion occurred by two distinct mechanisms, either by direct or by misalignment extension. An "A-rule" appeared to hold for the case of direct extension, where the abasic template moiety is intrahelical, aligned opposite the primer 3'-terminus. In misalignment extension, the primer terminus is realigned from a site directly opposite the lesion to a new position opposite a neighboring template base 5' to the lesion. Direct extension efficiencies were measured in 16 different configurations, by varying 4 bases at the primer 3'-termini and 4 at the 5'-side (downstream) of the lesion. The predominant order of direct extension was A > G > T approximately C, similar to that observed for insertion. Reduced primer extension rates were not caused by a reduction in HIV-1 RT-DNA binding. Primers terminating in C showed inefficient direct extension, but were readily extended via misaligned configurations. The ratios of direct-to-misalignment extension efficiencies were 27:1, 2.5:1, and 1:25 for A, G, and C opposite the lesion, respectively. For the case of primers terminating in T, misalignment extension was not observed. A striking result was that while primers were extended past an abasic lesion by HIV-1 RT in both direct and misalignment modes, avian myeloblastosis virus RT failed to catalyze significant extension by either mode.
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PMID:Kinetics of deoxyribonucleotide insertion and extension at abasic template lesions in different sequence contexts using HIV-1 reverse transcriptase. 769 91

Retroviral proteinase(PR)-catalyzed cleavage of the viral Gag and Gag-Pol polyproteins within the nascent virus particle is required for productive viral infection. Kinetic characterization and specificity analyses have been reported for several retroviral PR using oligopeptide substrates. In this study, we performed a comparative analysis of PR from avian, bovine, simian and human retroviruses using polyproteins of human immunodeficiency virus (HIV) type 1 or avian leukosis virus as substrates. Polyproteins were derived from immature virus-like particles purified from culture medium of transfected or recombinant baculovirus-infected cells. Specific cleavage to the correct size intermediate and end products occurred in the presence of detergent and homologous PR. HIV-1 PR cleaved its Gag precursor to completion at a concentration of approximately 25 nM but cleaved the Gag-Pol precursor incompletely even at fourfold higher PR concentration. In contrast to the requirement for high ionic strength for peptide cleavage reported previously, we found that Gag protein cleavage by HIV-1 PR proceeded best at low ionic strength, for both of the protein substrates tested. HIV-2 PR was approximately sixfold less active than HIV-1 PR. PR from avian myeloblastosis-associated virus (MAV) yielded efficient cleavage of the HIV-1 polyprotein only at concentrations above 1 microM. Both enzymes were stimulated by high salt and their cleavage products were identical or very similar to those of HIV-1 PR. A mutant of MAV PR engineered to cleave HIV-1 peptide substrates did not cleave the HIV-1 polyprotein at a concentration of 0.4 microM. The PR of Mason Pfizer monkey virus cleaved this polyprotein very poorly, whereas PR of bovine leukemia virus cleaved it, albeit at different sites.
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PMID:Proteolytic processing of particle-associated retroviral polyproteins by homologous and heterologous viral proteinases. 788 3

A steady state kinetic analysis of the avian myeloblastosis virus/Rous sarcoma virus (AMV/RSV) and human immunodeficiency virus Type 1 (HIV-1) retroviral proteases (PRs) was carried out using a series of 40 peptide substrates that are derivatives of the AMV/RSV nucleocapsid-PR cleavage site. These peptides contain single amino acid substitutions in each of the seven positions of the minimum length substrate required by the PR for specific and efficient cleavage. These peptide substrates are distinguished by the individual enzyme subsites of the AMV/RSV and HIV-1 PRs. The molecular basis for similarities and differences of the individual subsites for both proteases is discussed using steady state kinetic data and modeling based on crystal structures.
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PMID:Comparison of the substrate-binding pockets of the Rous sarcoma virus and human immunodeficiency virus type 1 proteases. 838 61

O-4'-nor-2', 3'-dideoxy-2', 3'-didehydronucleoside 5'-triphosphates are shown to be effective termination substrates of DNA biosynthesis catalyzed by human placental DNA polymerases alpha and epsilon, rat liver DNA polymerase beta, reverse transcriptases of human immunodeficiency virus and avian myeloblastosis virus, and calf thymus terminal deoxynucleotidyl transferase. These compounds do not interact only with the Escherichia coli DNA polymerase I (Klenow fragment). The probable reasons of interaction of acyclo-d4NTP with the DNA synthesizing complexes are discussed.
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PMID:[Acyclic analogs of 2',3'-dideoxy-2',3'-didehydronucleoside-5'-triphosphates--terminators of DNA synthesis, catalyzed by a broad set of DNA polymerases]. 848 66

The comparison of Km and Vmax values for various primers in the reaction of polymerization catalyzed by the human immunodeficiency virus type-1 (HIV-1) reverse transcriptase was carried out. The primers were: (a) complementary to the template, (b) partially complementary with mismatched nucleotides at different positions from the 3' end or (c) non-complementary. Non-complementary primers were not elongated by HIV-1 reverse transcriptase. However, if they contained only one residue complementary to the template or an abasic unit at the 3' end, they could serve as primers. The most effective discrimination between matched and mismatched primers, due to a decrease in the affinity and Vmax, was found in the case of oligonucleotides containing non-complementary bases at the second or third position from the 3' end of the primer. The efficiency of discrimination by HIV-1 reverse transcriptase between matched and mismatched base-paired primers was about 1-1.5 orders of magnitude lower than that of procaryotic, eucaryotic and archaebacterial DNA polymerases and avian myeloblastosis virus reverse transcriptase. Oligonucleotides such as (dT)4(dCdG)k(dT)4 showed higher affinity for the enzyme than (dT)4 or (dT)8 primers. These data suggest that HIV-1 reverse transcriptase, in contrast to procaryotic, eucaryotic and archaebacterial DNA polymerases, forms additional contacts with the 5'-end region of the non-complementary primer. In addition, using tRNA(3Lys), the natural primer of HIV-1, it was shown that the p66 subunit of reverse transcriptase can be crosslinked, in the presence of a platinum derivative, to the 5' end of tRNA. Thus, besides the normal binding site for the 3' end of tRNA, which is crucial for the initiation of cDNA synthesis, the 5' end of the tRNA also interacts with a specific site on the enzyme.
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PMID:High-affinity interaction of human immunodeficiency virus type-1 reverse transcriptase with partially complementary primers. 852 51

RNA hypermutagenesis results from cDNA synthesis in the presence of highly biased dNTP precursor concentrations and preferentially exploits human immunodeficiency virus type 1 (HIV-1) reverse transcriptase. Such reaction conditions slow down DNA synthesis, which might be conducive to strand transfer and deletion. This has been investigated. A 6 bp inverted repeat nested between 10 bp repeats was efficiently deleted at dCTP concentrations typically used. Inter- or intramolecular strand transfer between 10 bp repeated sequences separated by runs of templated G residues occurred, but at lower concentrations. If RNA hypermutagenesis of a sequence containing direct and inverted repeats is unavoidable, avian myeloblastosis virus (AMV) reverse transcriptase could be used, as strand transfer occurs with much diminished dCTP substrate dependence.
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PMID:Fate of direct and inverted repeats in the RNA hypermutagenesis reaction. 862 47

The 3SR (self-sustained sequence-replication) reaction is a very efficient method for isothermal amplification of target DNA or RNA sequences in vitro. This method requires three enzymatic activities: reverse transcriptase, DNA-dependent RNA polymerase and Escherichia coli ribonuclease H. We have modified the original protocol by using human immunodeficiency virus (HIV)-1 reverse transcriptase instead of avian myeloblastosis virus (AMV) reverse transcriptase to allow amplification with T7 RNA polymerase but without E. coli ribonuclease H. Comparison of the incorporation kinetics between the conventional three-enzyme 3SR and our two-enzyme 3SR shows differences in the kinetic behaviour. Furthermore, by the new two-enzyme 3SR, the amplified RNA is obtained in a purer form compared with the experiments with three-enzyme 3SR. The aim of our research is to adapt 3SR as a useful tool for darwinian evolutionary experiments.
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PMID:Comparison of self-sustained sequence-replication reaction systems. 863 38

The retroviral proteinase (PR) seems to play crucial roles in the viral life cycle, therefore it is an attractive target for chemotherapy. Previously we studied the specificity of human immunodeficiency virus (HIV) type 1 and type 2 as well as equine infectious anemia virus PRs using oligopeptide substrates. Here a similar approach is used to characterize the specificity of avian myeloblastosis virus (AMV) PR and to compare it with those of the previously characterized lentiviral PRs. All peptides representing naturally occurring Gag and Gag-Pol cleavage sites were substrates of the AMV PR. Only half of these peptides were substrates of HIV-1 PR. The Km values for AMV PR were in a micromolar range previously found for the lentiviral PRs; however, the kcat values were in a 10 30-fold lower range. A series of peptides containing single amino acid substitutions in a sequence representing a naturally occurring HIV cleavage site was used to characterize the seven substrate binding subsites of the AMV PR. The largest differences were found at the P4 and P2 positions of the substrate. Detailed analysis of the results by molecular modeling and comparison with previously reported data revealed the common characteristics of the specificity of the retroviral PRs as well as its strong dependence on the sequence context of the substrate.
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PMID:Comparative studies on the substrate specificity of avian myeloblastosis virus proteinase and lentiviral proteinases. 863

A non-radioactive 96-well microtitre plate reverse transcriptase (RT) assay, based on the use of covalently bound riboadenosine homopolymer in the wells and 5-bromodeoxyuridined 5'-triphosphate (BrdUTP) as dNTP, is described. The whole assay is performed in a single well, including the quantitative detection of incorporated BrdU, which is performed immunologically using alkaline phosphatase-conjugated anti-BrdU antibody and colorometric reading. The system also allows the use of variable amounts of primer. The kinetics and characteristics of the assay using BrdUTP is similar to the use of [3H]dTTP. The sensitivity of the assay can be varied either by altering the duration of RT assay time and/or by prolonging the alkaline phosphatase reaction. Thus the assay can detect < 0.02 pg of recombinant human-immunodeficiency-virus (HIV) type I RT, < 0.005 m unit of avian-myeloblastosis-virus RT or < 0.02 m unit of recombinant Moloney-murine-leukaemia-virus RT. The assay was found to be useful with various types of cell-culture material, and a comparative study of 16 HIV-infected lymphocyte cultures, using 10 microliters of supernatant medium for RT assay and 22.5 microliters for p24 antigen assay showed that the new RT assay was at least 25-fold more sensitive than the p24 antigen assay. The results also show a good correlation between the RT activities found and the p24-antigen level detected, with exception for HIV2 isolates, as they only became positive in the RT assay. The technical performance and the capacity of the test compared with other available RT kits is discussed, as well as its use for other applications.
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PMID:A sensitive assay for the quantification of reverse transcriptase activity based on the use of carrier-bound template and non-radioactive-product detection, with special reference to human-immunodeficiency-virus isolation. 863 77

High-affinity, high-specificity RNA ligands for reverse transcriptase from feline immunodeficiency virus (FIV) were isolated from an RNA library by the SELEX (Systematic Evolution of Ligands by EXponential enrichment) procedure. The selected RNA ligands bound to FIV reverse transcriptase with dissociation constants in the nanomolar range. One of the ligands was a potent inhibitor of the RNA-dependent DNA polymerase activity of both the recombinant and the virion-derived FIV reverse transcriptase. It also inhibited the reverse transcriptase from an FIV mutant that is resistant to 3'-azido-3'-deoxythymidine (AZT). The inhibition of FIV reverse transcriptase was competitive with respect to template-primer and noncompetitive with respect to deoxyribonucleoside 5'-triphosphates. This ligand was specific for the FIV enzyme and did not inhibit other reverse transcriptases tested (avian myeloblastosis virus, Moloney murine leukemia virus, and human immunodeficiency virus type 1).
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PMID:Inhibitory RNA ligand to reverse transcriptase from feline immunodeficiency virus. 863 44


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