Full Peptide Synthesis, Purification, and Characterization of Six Tat Variants

Abstract

AIDS in Africa is characterized by the equal distribu- tion of mortality between the two genders because of highly virulent human immunodeficiency virus type 1 (HIV-1) strains. The viral protein Tat trans-activates vi- ral gene expression and is essential for HIV-1 replica- tion. We chemically synthesized six different Tat pro- teins, with sizes ranging from 86 to 101 residues, from HIV-1 isolates located in different parts of the world including highly virulent African strains. Protein puri- fication, mass spectroscopy, and amino acid analysis showed that the synthesis was successful in each case but with different yields. We show that all have the ability to bind the HIV long terminal repeat (LTR) RNA trans-activation response element (TAR) region, in- volved in Tat-mediated trans-activation, but structural heterogeneities are revealed by circular dichroism. These Tat synthetic proteins cross membranes but differ in their ability to trans-activate an HIV LTR-reporter gene in stably transfected HeLa cells. Two Tat proteins from virulent African HIV-1 strains were much more active than those from Europe and the United States. The interferon-induced kinase (PKR), involved in cell antiviral defense, phosphorylates only Tat variants cor- responding to less or nonvirulent HIV-1 isolates. Our results indicate that the high virulence of some African HIV-1 strains could be related to Tat activity.