PROBING THE STRUCTURE OF THE HIV-1 REV TRANSACTIVATOR PROTEIN BY FUNCTIONAL-ANALYSIS

Human immunodeficiency virus type 1 (HIV-1) encodes a trans-acting reg ulatory protein, termed Rev, which is critically required for virus re plication. Rev is a sequence-specific RNA binding protein which mediat es the nuclear export of unspliced and incompletely spliced viral mRNA s encoding the viral structural proteins. While CD and fluorescence me asurements have provided several possible structural models of Rev, al l attempts employing X-ray crystallography and MMR techniques have so far failed to provide more accurate data. We present a new approach to validate alternative structural models of the N-terminal region of Re v which contains the nuclear localization/RNA binding domain. Points o f contact between structural elements in a protein were determined by introduction of targeted amino acid substitutions and subsequent scori ng of the biological activities. Our data resulted in the suggestion o f a new and more refined model of HIV-1 Rev structure which to date ha s been impossible to obtain by other means.