High resolution X-ray structure of potent anti-HIV pokeweed antiviral protein-III.
Pokeweed antiviral protein III (PAP-III), a naturally occurring protein isolated from late summer leaves of the pokeweed plant (Phytolacca americana), has potent anti-HIV activity by an as yet undetermined molecular mechanism. PAP-III belongs to a family of ribosome-inactivating proteins that catalytically deadenylate ribosomal and viral RNA. The chemical modification of PAP-III by reductive methylation of its lysine residues significantly improved the crystal quality for X-ray diffraction studies. Trigonal crystals of the modified PAP-III, with unit cell parameters a=b=80.47A, c=76.21A, were obtained using 30% PEG400 as the precipitant. These crystals contained one enzyme molecule per asymmetric unit and diffracted up to 1.5A, when exposed to a synchrotron source. Here we report the X-ray crystal structure of PAP-III at 1.6A resolution, which was solved by molecular replacement using the homology model of PAP-III as a search model. The fold typical of other ribosome-inactivating proteins is conserved, despite several differences on the surface and in the loop regions. Residues Tyr(69), Tyr(117), Glu(172), and Arg(175) are expected to define the active site of PAP-III. Molecular modeling studies of the interactions of PAP-III and PAP-I with a single-stranded RNA heptamer predicted a more potent anti-HIV activity for PAP-III due to its unique surface topology and more favorable charge distribution in its 20A-long RNA binding active center cleft. In accordance with the predictions of the modeling studies, PAP-III was more potent than PAP-I in depurinating HIV-1 RNA.
Kurinov
Igor V
IV
The Biotherapy and Drug Discovery Program, Parker Hughes Cancer Center, 2699 Patton Road, St. Paul, MN 55113, USA.
Uckun
Fatih M
FM
D003460:Crystallization; D018360:Crystallography, X-Ray; D006678:HIV / Q000187:drug effects; D008958:Models, Molecular; D009699:N-Glycosyl Hydrolases; D010940:Plant Proteins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D011487:Protein Conformation; D012313:RNA / Q000378:metabolism; D054789:Ribosome Inactivating Proteins, Type 1; D012270:Ribosomes / Q000187:drug effects
D010940:Plant Proteins; D054789:Ribosome Inactivating Proteins, Type 1; D012313:RNA; D009699:N-Glycosyl Hydrolases; C024275:pokeweed antiviral protein