Fusion partner toolchest for the stabilization and crystallization of G protein-coupled receptors.
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.
Department of Molecular Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA.
Eugene
E
Chun
Aaron A
AA
Thompson
Wei
W
Liu
Christopher B
CB
Roth
Mark T
MT
Griffith
Vsevolod
V
Katritch
Joshua
J
Kunken
Fei
F
Xu
Vadim
V
Cherezov
Michael A
MA
Hanson
Raymond C
RC
Stevens
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D045303:Cytochromes b / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D006801:Humans; D008969:Molecular Sequence Data; D009113:Muramidase / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D055550:Protein Stability; D043705:Receptor, Adenosine A2A / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D043562:Receptors, G-Protein-Coupled / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification
D043705:Receptor, Adenosine A2A; D043562:Receptors, G-Protein-Coupled; D011993:Recombinant Fusion Proteins; D045303:Cytochromes b; D009113:Muramidase