Nicking activity on pBR322 DNA of ribosome inactivating proteins from Phytolacca dioica L. leaves.
Ribosome-inactivating proteins isolated from Phytolacca dioica L. leaves are rRNA-N-glycosidases, as well as adenine polynucleotide glycosylases. Here we report that some of them cleave supercoiled pBR322 dsDNA, generating relaxed and linear molecules. PD-L1, the glycosylated major form isolated from the winter leaves of adult P . dioica plants, produces both free 3'-OH and 5'-P termini randomly distributed along the DNA molecule, as suggested by labelling experiments with [alpha- 32P]dCTP and [gamma- 32 P]dATP. Moreover, when the reaction is carried out under low-salt conditions, cleavage is observed mainly at a specific site, located downstream of the ampicillin resistance gene (close to position 3200), ending with the deletion of a fragment of approximately 70 nucleotides. This cleavage pattern is similar to that obtained under the same conditions with mung bean nuclease, a single-strand endonuclease. Furthermore, pBR322 DNA treated with PD-L1 shows reduced transforming activity with E . coli HB101 competent cells in comparison to untreated control plasmid DNA.
Aceto
Serena
S
Dipartimento di Genetica, Biologia Generale e Molecolare, Università degli Studi di Napoli Federico II, Via Mezzocannone 8, I-80134 Napoli, Italy.
Di Maro
Antimo
A
Conforto
Barbara
B
Siniscalco
Gesualdo G
GG
Parente
Augusto
A
Delli Bovi
Pasquale
P
Gaudio
Luciano
L
D001483:Base Sequence / Q000235:genetics; D004278:DNA, Superhelical / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D029603:Phytolacca; D018515:Plant Leaves; D010940:Plant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010957:Plasmids; D012335:RNA, Ribosomal / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D054789:Ribosome Inactivating Proteins, Type 1
D004278:DNA, Superhelical; D010940:Plant Proteins; D012335:RNA, Ribosomal; D054789:Ribosome Inactivating Proteins, Type 1; D009699:N-Glycosyl Hydrolases; C024275:pokeweed antiviral protein