Pokeweed antiviral protein isoforms PAP-I, PAP-II, and PAP-III depurinate RNA of human immunodeficiency virus (HIV)-1

Pokeweed antiviral protein (PAP) is a naturally occurring broad-spectrum antiviral agent with potent anti-human immunodeficiency virus (HIV)-1 activity by an as yet undeciphered molecular mechanism. In the present study, we sought to determine if PAP is capable of recognizing and depurinating viral RNA. Depurination of viral RNA was monitored by directly measuring the amount of the adenine base released from the viral RNA species using quantitative high-performance liquid chromatography. Our findings presented herein provide direct evidence that three different PAP isoforms from Phytolacca americana (PAP-I from spring leaves, PAP-II from early summer leaves, and PAP-III from late summer leaves) cause concentration-dependent depurination of genomic RNA (63 to 400 pmols of adenine released per micrograms of RNA) purified from human immunodeficiency virus type-I (HIV-I), plant virus (tobacco mosaic virus (TMV), and bacteriophage (MS 2). In contrast to the three PAP isoforms, ricin A chain (RTA) failed to cause detectable depurination of viral RNA even at 5 microM, although it was as effective as PAP in inhibiting protein synthesis in cell-free translation assays. PAP-I, PAP-II, and PAP-III (but not RTA) inhibited the replication of HIV-1 in human peripheral blood mononuclear cells with IC(50) values of 17 nM, 25 nM, and 16 nM, respectively. These findings indicate that the highly conserved active site residues responsible for the depurination of rRNA by PAP or RTA are not sufficient for the recognition and depurination of viral RNA. Our study prompts the hypothesis that the potent antiviral activity of PAP may in part be due to its unique ability to extensively depurinate viral RNA, including HIV-1 RNA.