Efficient protein production inspired by how spiders make silk

Nat Commun. 2017 May 23:8:15504. doi: 10.1038/ncomms15504.

Abstract

Membrane proteins are targets of most available pharmaceuticals, but they are difficult to produce recombinantly, like many other aggregation-prone proteins. Spiders can produce silk proteins at huge concentrations by sequestering their aggregation-prone regions in micellar structures, where the very soluble N-terminal domain (NT) forms the shell. We hypothesize that fusion to NT could similarly solubilize non-spidroin proteins, and design a charge-reversed mutant (NT*) that is pH insensitive, stabilized and hypersoluble compared to wild-type NT. NT*-transmembrane protein fusions yield up to eight times more of soluble protein in Escherichia coli than fusions with several conventional tags. NT* enables transmembrane peptide purification to homogeneity without chromatography and manufacture of low-cost synthetic lung surfactant that works in an animal model of respiratory disease. NT* also allows efficient expression and purification of non-transmembrane proteins, which are otherwise refractory to recombinant production, and offers a new tool for reluctant proteins in general.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cholecystokinin / chemistry
  • Chromatography
  • Circular Dichroism
  • Dimerization
  • Disease Models, Animal
  • Escherichia coli / metabolism
  • Female
  • Fibroins / biosynthesis
  • Hydrogen-Ion Concentration
  • Lung / pathology
  • Magnetic Resonance Spectroscopy
  • Micelles
  • Microscopy, Electron, Transmission
  • Mutagenesis, Site-Directed
  • Mutation
  • Peptides / chemistry
  • Protein Domains
  • Rabbits
  • Recombinant Proteins / biosynthesis*
  • Respiration Disorders / drug therapy
  • Silk / biosynthesis*
  • Spiders
  • Surface-Active Agents / chemistry*

Substances

  • Micelles
  • Peptides
  • Recombinant Proteins
  • Silk
  • Surface-Active Agents
  • Fibroins
  • Cholecystokinin