Biosynthesis and post-translational processing of the precursor to brain-derived neurotrophic factor

J Biol Chem. 2001 Apr 20;276(16):12660-6. doi: 10.1074/jbc.M008104200. Epub 2001 Jan 10.

Abstract

We examined the biosynthesis and post-translational processing of the brain-derived neurotrophic factor precursor (pro-BDNF) in cells infected with a pro-BDNF-encoding vaccinia virus. Metabolic labeling, immunoprecipitation, and SDS-polyacrylamide gel electrophoresis reveal that pro-BDNF is generated as a 32-kDa precursor that is N-glycosylated and glycosulfated on a site, within the pro-domain. Some pro-BDNF is released extracellularly and is biologically active as demonstrated by its ability to mediate TrkB phosphorylation. The precursor undergoes N-terminal cleavage within the trans-Golgi network and/or immature secretory vesicles to generate mature BDNF (14 kDa). Small amounts of a 28-kDa protein that is immunoprecipitated with BDNF antibodies is also evident. This protein is generated in the endoplasmic reticulum through N-terminal cleavage of pro-BDNF at the Arg-Gly-Leu-Thr(57)- downward arrow-Ser-Leu site. Cleavage is abolished when Arg(54) is changed to Ala (R54A) by in vitro mutagenesis. Blocking generation of 28-kDa BDNF has no effect on the level of mature BDNF and blocking generation of mature BDNF with alpha(1)-PDX, an inhibitor of furin-like enzymes, does not lead to accumulation of the 28-kDa form. These data suggest that 28-kDa pro-BDNF is not an obligatory intermediate in the formation of the 14-kDa form in the constitutive secretory pathway.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Brain-Derived Neurotrophic Factor / biosynthesis*
  • Brain-Derived Neurotrophic Factor / genetics
  • Brain-Derived Neurotrophic Factor / metabolism*
  • Brain-Derived Neurotrophic Factor / pharmacology
  • COS Cells
  • Cell Line
  • Chlorocebus aethiops
  • Embryo, Mammalian
  • Glycoside Hydrolases
  • Glycosylation
  • Hippocampus / metabolism*
  • Humans
  • Mice
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Neuroglia / metabolism
  • Neurons / metabolism*
  • Phosphorylation
  • Protein Precursors / biosynthesis
  • Protein Precursors / genetics
  • Protein Precursors / metabolism*
  • Protein Processing, Post-Translational*
  • Receptor, trkB / drug effects
  • Receptor, trkB / physiology
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / metabolism
  • Transfection
  • Vaccinia virus / genetics

Substances

  • Brain-Derived Neurotrophic Factor
  • Protein Precursors
  • Recombinant Proteins
  • brain-derived neurotrophic factor precursor
  • Receptor, trkB
  • Glycoside Hydrolases