Purification and characterization of novel ribosome inactivating proteins, alpha- and beta-pisavins, from seeds of the garden pea Pisum sativum.
Two ribosome inactivating proteins designated alpha- and beta-pisavins were isolated from seeds of the garden pea Pisum sativum var. arvense Poir with a procedure involving affinity chromatography on Affi-gel Blue gel, immobilized metal ion affinity chromatography on Iminodiacetic acid-agarose, cation exchange chromatography on Resource-S, and gel filtration on Superose 12. alpha- and beta-pisavins are nonglycoproteins with a molecular weight of 20.5 kDa and 18.7 kDa respectively. The sequences of the first sixty N-terminal amino acids of alpha- and beta-pisavins were identical. In isoelectric focusing these two proteins merged into one band with a pI greater than 9.3. Inhibition of protein synthesis by a rabbit reticulocyte lysate system was achieved at an IC50 of approximately 0.5 nM. Activity of the proteins toward tRNA was observed. The proteins acted on ribosomal RNA through its RNA N-glycosidase activity to release an Endo's fragment, and converted the conformation of DNA from supercoiled and circular forms into a linear form.
Lam
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Department of Biochemistry, Faculty of Medicine, Chinese University of Hong Kong, People's Republic of China.
Wang
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Ng
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D000595:Amino Acid Sequence; D000818:Animals; D002474:Cell-Free System / Q000187:drug effects; D008969:Molecular Sequence Data; D008970:Molecular Weight; D018532:Pisum sativum / Q000737:chemistry*; D010446:Peptide Fragments / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D011500:Protein Synthesis Inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D011817:Rabbits; D012156:Reticulocytes; D012270:Ribosomes / Q000187:drug effects / Q000378:metabolism*; D012639:Seeds / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid
D010446:Peptide Fragments; D010940:Plant Proteins; D011500:Protein Synthesis Inhibitors; C116393:pisavin protein, Pisum sativum