Expression and characterisation in E. coli of mutant forms of saporin

J Biotechnol. 2005 May 25;117(3):263-6. doi: 10.1016/j.jbiotec.2005.01.021. Epub 2005 Apr 9.

Abstract

In the present communication, we report on the expression and characterisation in Escherichia coli of mutant derivatives of saporin, a type 1 ribosome-inactivating protein from Saponaria officinalis L. The effects of substitution of Glu 176 with Lys and those of deletion of 19 amino acids at the C-terminal were evaluated both in vivo, testing the influence of expressed proteins on bacterial growth and in vitro measuring their N-glycosidase and supercoiled DNA relaxation activities. Results indicate that both modifications of the wild-type protein abolish its toxicity to bacterial cells and impair its enzymatic activity on polynucleotide substrates, either RNA or DNA.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Amino Acid Substitution
  • Cell-Free System
  • DNA, Superhelical / metabolism
  • Escherichia coli / enzymology*
  • Escherichia coli / genetics
  • Escherichia coli / growth & development
  • Escherichia coli / metabolism*
  • Lysine / metabolism
  • Mutation*
  • N-Glycosyl Hydrolases / analysis
  • N-Glycosyl Hydrolases / metabolism
  • Plant Proteins / genetics*
  • Plant Proteins / toxicity*
  • Recombinant Proteins / toxicity
  • Saponaria / chemistry
  • Sequence Deletion

Substances

  • DNA, Superhelical
  • Plant Proteins
  • Recombinant Proteins
  • N-Glycosyl Hydrolases
  • Lysine