Enzyme reverse transcriptase

RNA oncogenic vimses have an unusual enzyme, reverse transcriptase, which is capable of making DNA copies from an RNA template. Cells transformed by these retrovimses have been shown to possess DNA transcripts of the viral RNA. It appears that the transformahon from normal to malignant is associated with the acquisition by the cell of viral DNA. 

A solution to the problem of introns is to isolate mRNA extracted from the human pancreas cells that make insulin. These cells are rich in insulin mRNA from which introns have already been spliced out. Using the enzyme reverse transcriptase it is possible to convert this spliced mRNA into a DNA copy. This copy DNA (cDNA), which carries the uninterrupted genetic information for insulin can be cloned. Although yeast cells (Saccharomyces) can splice out introns it is normal practice to eliminate them anyway by cDNA cloning. 

An extension of the PCR method is RT-PCR. Here mRNA is extracted from the cells or tissue, converted to cDNA using the enzyme reverse transcriptase and PCR is then carried out This method is used to detect the expression of specific mRNA sequences in cells or tissues. 

The enzyme reverse transcriptase, as well as some other enzymes used in virus replication the gene that codes these enzymes is the pol gene. The other viral enzymes specified by the pol gene are protease and integrase. Protease is involved in muturation of viral proteins as the virus buds out from the cell, and integrase is responsible for integration of the viral DNA into the cell s chromosomal DNA. 

Figure 20.18 The central dogma of molecular biology a summary of processes involved inflow of genetic information from DNA to protein. The diagram is a summary of the biochemical processes involved in the flow of genetic information from DNA to protein via RNA intermediates. This concept had to be revised following the discovery of the enzyme, reverse transcriptase, which catalyses information transfer from RNA to DNA (see Chapter 18). It may have to be modified in the future since changes in the fatty acid composition of phospholipids in membranes can modily the properties of proteins, and possibly their functions, independent of the genetic information within the amino acid sequence of the protein (See Chapters 7, 11 and 14).

The life history of a retroviras is described in chapter 17 (see Figure 17.45). A summary is presented here. The genome of a retrovirus is composed of RNA not DNA but, when a retrovirus infects a host cell its RNA is transcribed into DNA, catalysed by the enzyme, reverse transcriptase. This DNA is then incorporated into the genome of the host. On transcription of the host DNA, during cell division, viral mRNA and viral genomic RNA are produced. The... 

The virus that causes AIDS, the Human Immune Deficiency Virus (HIV) is a retrovirus. Instead of double-stranded DNA it uses single-stranded RNA to store its genetic information. HIV uses the enzyme reverse transcriptase to convert its RNA into DNA in order to replicate. 

The term oncogene was coined in association with the search for the tumor-causing principle in retroviruses. Retroviruses contain RNA as the genetic material and can transcribe RNA into DNA with the help of the virus s own enzyme reverse transcriptase. The DNA form of retroviruses can integrate into the DNA of the host cell and, during cell division, is passed on to the daughter cells as a provirus. From the provirus, viral RNA and complete virus particles may be formed. 

A limitation of this strategy is the fidelity of enzymes—reverse transcriptase and DNA polymerase— used in PCR to produce consistently error-free DNA sequences. These enzymes typically introduce error at the rate of one base per 400... 

Like all other retroviruses, human immunodeficiency virus type 1 (HIV-1) contains the multifunctional enzyme reverse transcriptase (RT). Retroviral RTs have a DNA polymerase activity that can use either an RNA or a DNA template and an RNase H activity. HIV-1 RT is essential for the conversion of single-stranded viral RNA into a linear double-stranded DNA that is subsequently integrated into the host cell chromosomes [1-4]. In this conversion process HIV-1 RT catalyzes the incorporation of approximately... 

Persistent viral infection is a difficult challenge for antiviral chemotherapy. Retroviruses as a class are often found to be responsible for persistent viral infections. Retroviruses are unique RNA viruses characterized by the transcription of their single-stranded RNA into the double-stranded DNA of the host cell using the viral enzyme reverse transcriptase. AIDS is an example of such a persistent and latent human viral infection. 

HIV is an RNA virus whose hallmark is the reverse transcription of its genomic RNA to DNA by the enzyme reverse transcriptase. The life cycle of HTV begins with the high-affinity binding of the gpl20 protein via a portion of its VI region near the N terminus to its receptor on the host cell surface, the CD4 molecule (Figure 40.1). 

The enzyme reverse transcriptase makes DNA from an KNA model—the reverse of the usual process. In this way, KNA viruses (retroviruses) can be incorporated into the chromosome as DNA copies. There is now evidence that DNA can be moved from one place to another in the genome by this method the DNA is transcribed to KNA, which in turn is transcribed to chromosomal DNA. Roughly, the consequences are the same as those of a transposon. 

The enzymes that manipulate nucleotides, nucleic acids, etc. are the points of therapeutic intervention for a number of diseases involving cell replication disorders such as cancers and viral infections. For instance, AZT 10.10, an inhibitor of the enzyme reverse transcriptase, is an anti-viral drug currently used in the treatment of AIDS. 

Proteins never serve as templates for RNA. However, RNA chains can in rare circumstances reverse the flow of information from RNA to DNA. An example of this is the infecting RNA of retroviruses that serves as a template for the synthesis of a single-stranded complementary DNA (cDNA) chain that functions as a template for a complementary DNA chain. The resulting double-stranded DNA then acts as a template for the synthesis of the original viral RNA chain. The virus-specific enzyme, reverse transcriptase, catalyzes the synthesis of DNA on the RNA template. Little if any reverse transcriptase activity is present in normal cells, so that very little DNA is formed on RNA templates. 

Human immunodeficiency virus (HIV) is a member of the retrovirus family, classified under the lentivirus genus. Retroviruses are enveloped RNA viruses, which contain a core of capsid proteins, viral RNA, and enzymes. All infectious retroviral virions contain an enzyme, reverse transcriptase, which catalyzes the formation of a complementary DNA strand from an RNA template. A double-stranded DNA copy of the viral RNA genome (proviral DNA) may then be integrated into and replicated with the host cell genome. 

When AIDS (Acquired Immune Deficiency Syndrome) first came into the news in the 1980s it was a horror story of mysterious deaths from normally harmless diseases after the patient s immune system had been weakened and eventually destroyed. The cause was identified by biologists as a new virus HIV (Human Immunodeficiency Virus) and antiviral drugs, notably AZT (Chapter 49), were used with some success. These drugs imitate natural nucleosides (AZT imitates deoxythymidine) and inhibit the virus from copying its RNA into DNA inside human cells by inhibiting the enzyme reverse transcriptase . 

Reverse transcription ofmRNA The use of the enzyme reverse transcriptase (RT) transcribes isolated mRNA into cDNA (copy DNA). 

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