Individual viral proteins

Despite these theoretical concerns, a number of HIV vaccines are under development. Most of these vaccines have been developed by recombinant DNA techniques that have allowed a large-scale production of individual viral proteins. The predominant HIV proteins that make up these potential vaccines are env proteins (e.g., gp 120) and, to a lesser extent, gag proteins. In addition, inactivated whole HIV virus is being tested. 

Once the viral RNA has been translated and the large polyprotein has been processed to form individual viral proteins, new viral particles begin to form. Packaged in each virion are two copies of fully unspliced viral RNA. To be packaged into the virion, the RNA dimerizes in a highly orchestrated process involving a self-complementary stem-loop interaction. Such HIV-specific RNA events may prove useful for fumre therapeutic intervention, although they have so far received relatively little attention by medicinal chemists. 

This approach was used to determine the complete antigen-specific humoral immune-response profile Ifom infected humans and animals. The vaccinia virus proteome containing 185 individual viral proteins was printed on a chip after cloning and expression. The chips were then used to determine the antibody profile in serum from vaccinia-virus-immunized humans, primates, and mice [113]. 

This test depends on attaching virus protein to a small laboratory dish. A serum sample is prepared from the blood of the individual to be tested, and it is placed in the dish containing bound HIV viral proteins. If HIV-specific antibodies are present in the serum, they will become tightly bound to the dish by way of the HIV proteins. The serum is then removed, and the dish is washed during this procedure, only antibodies specific for HIV will be retained. The dish is then reacted with a stain that will detect any human antibodies. Thus, dishes that were exposed to serum containing HIV-specific antibodies will be stained, while dishes from antibody-negative serum samples will be unstained. A modified ELISA test was developed in which the virus proteins are attached to small beads that can float in solution, instead of to the bottom of the dish. The test is carried out in a test tube and proceeds as before. 

Pharmacology Indinavir is an inhibitor of the HIV protease. HIV protease is an enzyme required for the proteolytic cleavage of the viral polyprotein precursors into the individual functional proteins found in infectious HIV. Indinavir binds to the protease active site and inhibits the activity of the enzyme. This inhibition prevents cleavage of the viral polyproteins resulting in the formation of immature... 

It is an inhibitor of the enzyme HIV protease which is required for the proteolytic cleavage of the viral polyprotein precursors into the individual functional proteins found in infectious HIV. 

It has been demonstrated that the chemical steps that comprise DNA integration are carried out by the viral protein, integrase (IN). Integrase is encoded by the 3 end of the viral pol gene, which also codes for two other viral enzymes, the protease and reverse transcriptase. These three enzymes are initially synthesized as part of a larger polyprotein that is subsequently cleaved by the protease to the individual proteins. 

Fig. I. Examples of Western blots stained with colloidal gold for total protein followed by immunostaining of individual antigensJ.anes 1—3 Proteins on a Western blot from a cytoplasmic extract of poliovirus-infected HEp-2 cells were stained with colloidal gold. The probing monoclonal antibodies, recognizing the viral proteins VP1 and precursor, VPO and VP2, and VP3, respectively, are detected by peroxidase-coupled rabbit-antimouse antibody (asterisks). Lane 4 Western blot of an E. coli lysate, containing a fusion protein composed of protein A and the poliovirus protein 2B. The fusion protein (arrowhead) is detected on the gold-stained blot by peroxidase-coupled IgG that binds to the protein A moiety.

Indinavir (800 mg/t.i.d.) is an inhibitor of the HIV protease, which is an enzyme required for the proteolytic cleavage of the viral polyprotein precursors into the individual functional proteins found in infectious HIV. Indinavir binds to the protease active site and inhibits the activity of the enzyme. This inhibition prevents cleavage of the viral polyproteins resulting in the formation of immature noninfectious viral particles. Cross-resistance between indinavir and HIV reverse-transcriptase inhibitors is unlikely because the enzyme targets involved are different. Cross-resistance was noted between indinavir and the protease inhibitor ritonavir. Varying degrees of cross-resistance have been noted between indinavir and other HIV protease inhibitors. Indinavir is metabolized in the liver, and seven metabolites have been identified, and 20% of indinavir is excreted unchanged in the urine. 

In their model Euss and Koch (40) propose that after infection there is a non-specific reduction in the overall rate of polypeptide chain initiation. However, since each mEEA has its own intrinsic translational efficiency, not all mEEAs are affected to the same extent, and there is a differential reduction in the translation of individual host and viral mEEA species. In a somewhat different model, Carrasco and Smith (4I, 42) propose that upon contact of the virus with the cell membrane, a viral coat protein associates with the membrane, changing the normal monovalent-ion gradient. During the course of viral EEA translation into progeny coat protein, these proteins are also inserted into the membrane, and the membrane alterations continue. Eventually, sodiim leaks into the cell, there is an increase in the concentration of monovalent-ions inside the cells, and cellular but not viral protein chain initiation is inhibited. 

Another icosahedral plant virus used as a viral template is CCMV, a member of the Bromoviridae family. CCMV has a diameter of 28.6 nm and is comprised of a coat protein shell encapsulating a single strand of positive-sense RNA. Similar to CPMV, the genome of CCMV is comprised of multiple strands of RNA, with the three unique strands of RNA packaged individually into virus particles. The individual coat protein capsids of CCMV are composed of 190 amino acid residues and have a total mass of 19.8 kDa. Each coat protein subunit folds into the canonical virus -barrel structure. The protein shell of the virus is composed of 180 identical coat protein subunits. In contrast to CPMV, CCMV particles are stable at pH 5 but swell up to 10% in size when the pH is increased to 7. As illustrated in Figure 7, when the virus is swollen, pores are created within the protein shell, which permit the interior of the virns particles to be penetrated by small molecnles. The transition... 

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