Purification and characterization of Moschatin, a novel type I ribosome-inactivating protein from the mature seeds of pumpkin (Cucurbita moschata), and preparation of its immunotoxin against human melanoma cells

Cell Res. 2003 Oct;13(5):369-74. doi: 10.1038/sj.cr.7290182.

Abstract

A novel ribosome-inactivating protein designated Moschatin from the mature seeds of pumpkin (Cucurbita moschata) has been successively purified to homogeneity, using ammonium sulfate precipitation, CM-cellulose 52 column chromatography, Blue Sepharose CL-6B Affinity column chromatography and FPLC size-exclusion column chromatography. Moschatin is a type 1 RIP with a pI of 9.4 and molecular weight of approximately 29 kD. It is a rRNA N-glycosidase and potently blocked the protein synthesis in the rabbit reticulocyte lysate with a IC50 of 0.26 nM. Using the anti-human melanoma McAb Ng76, a novel immunotoxin Moschatin-Ng76 was prepared successfully and it efficiently inhibited the growth of targeted melanoma cells M21 with a IC50 of 0.04 nM, 1500 times lower than that of free Moschatin. The results implied that Moschatin could be used as a new potential anticancer agent.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antineoplastic Agents, Phytogenic / chemistry
  • Antineoplastic Agents, Phytogenic / isolation & purification*
  • Antineoplastic Agents, Phytogenic / pharmacology
  • Cell Line, Tumor
  • Cucurbita / chemistry*
  • Drug Screening Assays, Antitumor
  • Humans
  • Immunoglobulin G / immunology
  • Immunotoxins / chemistry*
  • Immunotoxins / immunology
  • Immunotoxins / isolation & purification
  • Immunotoxins / pharmacology*
  • Melanoma / drug therapy*
  • Molecular Weight
  • N-Glycosyl Hydrolases / antagonists & inhibitors
  • N-Glycosyl Hydrolases / biosynthesis
  • N-Glycosyl Hydrolases / chemistry
  • N-Glycosyl Hydrolases / pharmacology*
  • Rabbits
  • Ribosome Inactivating Proteins
  • Ribosomes / drug effects
  • Seeds / chemistry*

Substances

  • Antineoplastic Agents, Phytogenic
  • Immunoglobulin G
  • Immunotoxins
  • N-Glycosyl Hydrolases
  • Ribosome Inactivating Proteins