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November 27, 1998
How HIV Evades AZT
HHMI researchers have used x-ray crystallography to produce a three-dimensional picture of a key AIDS virus enzyme. The structure reveals details that clarify how HIV mutations render the
retrovirus resistant to antiviral drugs such as AZT.
Once HIV invades a human cell, reverse transcriptase (RT), one of three
enzymes used by HIV, reads the retrovirus's genetic material, RNA, and
synthesizes DNA, which codes for viral proteins that commandeer the infected
cell's protein-making machinery. In an article published in the November 27,
1998, issue of the journal
Science,
Stephen C. Harrison
, an HHMI
investigator at Children's Hospital in Boston, and his colleagues from
Harvard University, describe the active structure of RT.
In order to capture RT in an active form, Harrison knew that the enzyme
needed to be constrained, so he approached Harvard chemists, Gregory Verdine
and his postdoctoral fellow Huifang Huang, and asked them to tether RT to
DNA. The chemists modified both DNA and the RT protein so that the two would
be attached by a permanent chemical bond. "The really clever new trick that
helped us trap a catalytically active complex was this modified chemistry,"
Harrison says.
The first images of the structure of RT were produced in the early 1990s by
HHMI investigator
Thomas A. Steitz
at Yale University. Steitz's research
team used a catalytically-inactive form of the protein in their studies.
Steitz and others showed that RT and other members of the DNA polymerase
family of enzymes are composed of subunits that resemble fingers, a palm,
and a thumb. In the RT found in HIV, DNA lies across this hand, stretching
from where the wrist would be toward the metaphorical pinkie. The structure
shows that active RT looks like a slightly clenched hand, with the thumb
almost touching the finger nearest to it.
Most drugs that target RT mimic nucleotides. When the drugs are incorporated
into DNA strands in the place of normal nucleotides, the strands stop
growing, and this effectively stops viral replication. Mutations that confer
resistance to AZT and other nucleotide analogs occur in the pocket between
the thumb and forefinger where the nucleotide binds. The bending of the
fingers that is clearly visible in the newly solved structure clarifies how
mutations alter binding to the DNA nucleotide or to the nucleotide analog
antiviral drugs such as AZT. Harrison's group shows that these mutations
prevent the nucleotide analog drugs from binding to the enzyme. These
mutations do not prevent normal nucleotides from binding, however, which
allows RT to continue making DNA for the virus. "Because of very elegant
chemistry accomplished by Verdine and Huang, we've been able to trap an
informative complex that allows us to understand much better the resistance
mutations," Harrison says.
Harrison says he is also excited about the research because the structure of
the catalytically-active form of RT may reveal general principles about how
viral polymerases work.
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