Agouron Pharmaceuticals Inc. scientists have taken a majorfirst step toward the rational development of drugs to blockHIV-1 infection.

The La Jolla, Calif.-based company reported today in Sciencethat it has determined the three-dimensional structure of anenzyme critical for replication of the AIDS virus HIV-1.

Agouron is now using the structure to identify small moleculesthat inhibit activity of the enzyme, reverse transcriptase (RT).

RT is a key in HIV replication and infection. It converts thesingle-stranded HIV RNA genome into a double-stranded DNAmolecule that inserts itself into a patient's DNA and producescopies of the virus.

RT's large size limits the ability of scientists to carry out X-raycrystallography of it. Instead, the Agouron team opted to makecrystals of a section of the enzyme known as the RNase Hdomain.

RT contains two domains. One copies the viral RNA into DNA(polymerase) and the second removes the RNA template (RNaseH).

Agouron scientists have previously reported that partialinactivation of RNase H blocks HIV replication, suggesting thatRNase H is a good domain to target for drug development.

To produce the large amounts of pure RNase needed todetermine its structure, the Agouron team designed arecombinant expression system that churned out the RNasedomain fused to another protein, dihydrofolate reductase.

Cleavage of this fusion protein by HIV protease left a "tail" of14 amino acids, usually considered to be part of thepolymerase domain, attached to the RNase domain. Themolecule made crystals suitable for high-resolution X-raydiffraction and three-dimensional structure determination.

Within five months of growing the crystals, the Agouron teamhad the answer to RNase H's structure and clues about how tocontrol its activity.

Comparisons between the HIV RNase H and a well-characterized RNase H isolated from E. coli show that the HIVstructure resembles the E. coli enzyme very closely. One majordifference is that although the E. coli enzyme is active on itsown, Agouron's RNase H domain only clips RNA if it is bound tothe polymerase domain of RT.

Agouron scientists believe that in HIV, the polymerase andRNase domains have evolved into structures that aredependent each other to keep the HIV genes in the properposition for construction of the new DNA version.

In an accompanying article, Science cites Stuart Legriece ofCase Western Reserve University in Cleveland as having an HIVRNase H molecule that does not require the polymerase domainfor activity. Zuzana Hostomska, one of the authors of today'sreport, said that Agouron is confident its RNase H has theproper structure.

-- Cynthia Robbins-Roth, Ph.D. BioVenture View

(c) 1997 American Health Consultants. All rights reserved.