Polynucleotide: adenosine glycosidase activity of saporin-L1: effect on DNA, RNA and poly(A)

Abstract

The ribosome-inactivating proteins (RIPs) are a family of plant enzymes for which a unique activity has been determined: rRNA N-glycosidase, which removes adenine at a specific universally conserved position (A4324 in the case of rat ribosomes). Here we report that saporin-L1, a RIP from the leaves of Saponaria officinalis, recognizes other substrates, including RNAs from different sources, DNA and poly(A). Saporin-L1 depurinated DNA extensively and released adenine from all adenine-containing polynucleotides tested. Adenine was the only base released from DNA or artificial polynucleotides. The characteristics of the reactions catalysed by saporin-L1 have been determined: optimal pH and temperature, ionic requirements, and the kinetic parameters Km and kcat. The reaction proceeded without cofactors, at low ionic strength, in the absence of Mg2+ and K+. Saporin-L1 had no activity towards various adenine-containing non-polynucleotide compounds (cytokinins, cofactors, nucleotides). This plant protein may now be classified as a polynucleotide:adenosine glycosidase.

A systemic antiviral resistance‐inducing protein isolated from Clerodendrum inerme Gaertn. is a polynucleotide:adenosine glycosidase (ribosome‐inactivating protein)

“…These effects are catalytic, as shown by molar ratio less than 1 between RIP present in the reaction mixture and adenine released. From the characteristics of their enzymatic activity, RIPs appear to be a non-homogeneous class of enzymes, in that (i) some release one, and some more than one adenine residues per ribosome [14], (ii) some are more active on poly(A) than on RNA [15,16] and (iii) have variable activity on RNA and on DNA (unpublished results). CIP-29 releases only one adenine residue from rat liver ribosomes, is not particularly active on poly(A), but is very active on DNA, more than any RIP assayed so far under the same experimental conditions.…”

“…RIPs were found to be peculiar N-glycosidases, and to remove a single adenine from a precise position in rRNA (A 4324 in the case of rat liver ribosomes) (review in [13]). More recently, however, it was reported that some, but not all, RIPs remove more than one adenine residue per ribosome [14], and that saporins [15,16] and other RIPs ( [12]; unpublished results by Barbieri et al) remove adenine from RNA other than rRNA and also from DNA. Consistently, ricin removes 2'-deoxyadenine from a synthetic dodecanucleotide with a GdA-GA loop, actually at a faster rate than it removes adenine from a similar dodecanucleotide with a GAGA loop [17].…”

The N-Glycosidase Activity of the Ribosome-inactivating Protein ME1 Targets Single-stranded Regions of Nucleic Acids Independent of Sequence or Structural Motifs

“…For instance, it has been shown that several RIPs can release adenine from multiple sites in rRNA (5). Furthermore, saporin-L1 can release adenine residues from a variety of nucleic acid substrates, including poly(A), mRNA, tRNA, and DNA (6,7). More than 50 other RIPs are active on DNA (8).…”

Insights into the Molecular Antiviral Mechanism of Pokeweed Protein from Phytolacca americana

“…In addition to ribosomal depurination, some RIPs act on naked rRNA (rRNA devoid of ribosomal proteins); some remove multiple adenines and guanines from various nucleic acid substrates including DNA, poly(A), rRNA, and even viral RNA [29][30][31][32]. Viral RNA depurination is a matter that is worthy of attention.…”