Site-specific mutagenesis of mistletoe lectin: the role of RIP activity in apoptosis

Biochem Biophys Res Commun. 1999 Nov 2;264(3):944-8. doi: 10.1006/bbrc.1999.1610.


Recombinant mistletoe lectin (rML) belongs to the class of type II ribosome-inactivating proteins (RIP) composed of a catalytically active A-chain with rRNA N-glycosidase activity and a B-chain with carbohydrate binding properties. To investigate the contribution of the enzymatic activity of the rML A-chain to the observed cytotoxic and apoptotic effects, an rMLA E166Q R169Q molecule was developed by means of site-specific mutagenesis. Following heterologous expression, the activity of mutant rMLA was measured in a cell-free assay for rRNA-N-glycosidase activity. Moreover, after generation of heterodimer, the activities of mutant rML E166Q R169Q and rML wild type were determined in a cytotoxicity and apoptosis assay. Although the reduction of activity as measured in the cell-free RIP assay was more pronounced (factor 237) than in both cellular assays (factors 20-22), the data clearly indicate a close correlation between cytotoxicity, apoptosis, and the enzymatic activity of the rML A-chain. Thus, RIP activity is an essential feature of rML and therefore a prerequisite for its biological function as an anticancer agent.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis / genetics*
  • Lectins / genetics*
  • Lectins / metabolism
  • Mistletoe
  • Mutagenesis, Site-Directed
  • Plant Lectins
  • Plant Preparations*
  • Plant Proteins*
  • Plants, Medicinal
  • Protein Synthesis Inhibitors / metabolism
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Ribonucleosides / genetics
  • Ribonucleosides / metabolism
  • Ribosome Inactivating Proteins, Type 2
  • Ribosomes
  • Toxins, Biological / genetics*
  • Toxins, Biological / metabolism


  • Lectins
  • Plant Lectins
  • Plant Preparations
  • Plant Proteins
  • Protein Synthesis Inhibitors
  • Recombinant Proteins
  • Ribonucleosides
  • Ribosome Inactivating Proteins, Type 2
  • Toxins, Biological
  • ribosome inactivating protein, Viscum