Fusion partner toolchest for the stabilization and crystallization of G protein-coupled receptors

Structure. 2012 Jun 6;20(6):967-76. doi: 10.1016/j.str.2012.04.010.

Abstract

Structural studies of human G protein-coupled receptors (GPCRs) have recently been accelerated through the use of a fusion partner that was inserted into the third intracellular loop. Using chimeras of the human β(2)-adrenergic and human A(2A) adenosine receptors, we present the methodology and data for the initial selection of an expanded set of fusion partners for crystallizing GPCRs. In particular, use of the thermostabilized apocytochrome b(562)RIL as a fusion partner displays certain advantages over previously utilized fusion proteins, resulting in a significant improvement in stability and structure of GPCR-fusion constructs.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cell Line
  • Chromatography, Gel
  • Cloning, Molecular
  • Crystallization
  • Crystallography, X-Ray
  • Cytochromes b / biosynthesis
  • Cytochromes b / chemistry*
  • Cytochromes b / isolation & purification
  • Humans
  • Molecular Sequence Data
  • Muramidase / biosynthesis
  • Muramidase / chemistry*
  • Muramidase / isolation & purification
  • Protein Stability
  • Receptor, Adenosine A2A / biosynthesis
  • Receptor, Adenosine A2A / chemistry*
  • Receptor, Adenosine A2A / isolation & purification
  • Receptors, G-Protein-Coupled / biosynthesis
  • Receptors, G-Protein-Coupled / chemistry
  • Receptors, G-Protein-Coupled / isolation & purification
  • Recombinant Fusion Proteins / biosynthesis
  • Recombinant Fusion Proteins / chemistry*
  • Recombinant Fusion Proteins / isolation & purification

Substances

  • Receptor, Adenosine A2A
  • Receptors, G-Protein-Coupled
  • Recombinant Fusion Proteins
  • Cytochromes b
  • Muramidase