A new approach to artificial and modified proteins: theory-based design, synthesis in a cell-free system and fast testing of structural properties by radiolabels

Protein Eng. 1994 Aug;7(8):1041-52. doi: 10.1093/protein/7.8.1041.

Abstract

A novel approach to the creation of artificial and modified proteins has been elaborated. The approach includes a sequence design based on the molecular theory of protein secondary structure and folding patterns, gene expression in a cell-free system and testing of structural properties of the synthesized polypeptides at a nanogram level using radiolabelled chains. The approach has been applied to a new synthetic protein albebetin which has been designed to form a 3-D fold which does not contradict any structural rule but has been never observed up to now in natural proteins. Using size-exclusion chromatography, urea-gradient electrophoresis and limited proteolysis of a radiolabelled chain, it has been shown that the artificial protein is nearly as compact as natural proteins, cooperatively unfolds at high urea concentrations and has some structural features of a definite structure consistent with the designed one. As albebetin has been designed as consisting of two structural repeats, a 'half-albebetin' (one of these repeats) has also been synthesized and studied. It was shown that 'half-albebetin' is also compact.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Chemical Phenomena
  • Chemistry, Physical
  • Genes, Synthetic
  • Isotope Labeling
  • Models, Molecular
  • Molecular Sequence Data
  • Peptide Fragments / chemical synthesis
  • Protein Biosynthesis
  • Protein Denaturation
  • Protein Engineering / methods*
  • Protein Folding*
  • Protein Structure, Secondary*
  • Protein Structure, Tertiary
  • Proteins / chemistry*
  • Proteins / drug effects
  • Proteins / genetics
  • Transcription, Genetic
  • Urea / pharmacology

Substances

  • Peptide Fragments
  • Proteins
  • albebetin
  • Urea