2K9P

Structure of TM1_TM2 in LPPG micelles

Summary for 2K9P

• Entry DOI: 10.2210/pdb2k9p/pdb
• NMR Information: BMRB: 15995
• Descriptor: Pheromone alpha factor receptor (1 entity in total)
• Functional Keywords: gpcr, micelle, structurral biology, fragment, g-protein coupled receptor, glycoprotein, membrane, pheromone response, phosphoprotein, receptor, transducer, transmembrane, membrane protein
• Biological source: Saccharomyces cerevisiae (yeast)
• Total number of polymer chains: 1
• Total formula weight: 8757.15

Authors

Neumoin, N.,Zerbe, O.,Naider, F. (deposition date: 2008-10-21, release date: 2009-05-05, Last modification date: 2024-05-08)

Primary citation

Neumoin, A.,Cohen, L.S.,Arshava, B.,Tantry, S.,Becker, J.M.,Zerbe, O.,Naider, F.
Structure of a double transmembrane fragment of a G-protein-coupled receptor in micelles.
Biophys.J., 96:3187-3196, 2009

PubMed Abstract: The structure and dynamic properties of an 80-residue fragment of Ste2p, the G-protein-coupled receptor for alpha-factor of Saccharomyces cerevisiae, was studied in LPPG micelles with the use of solution NMR spectroscopy. The fragment Ste2p(G31-T110) (TM1-TM2) consisted of 19 residues from the N-terminal domain, the first TM helix (TM1), the first cytoplasmic loop, the second TM helix (TM2), and seven residues from the first extracellular loop. Multidimensional NMR experiments on [(15)N], [(15)N, (13)C], [(15)N, (13)C, (2)H]-labeled TM1-TM2 and on protein fragments selectively labeled at specific amino acid residues or protonated at selected methyl groups resulted in >95% assignment of backbone and side-chain nuclei. The NMR investigation revealed the secondary structure of specific residues of TM1-TM2. TALOS constraints and NOE connectivities were used to calculate a structure for TM1-TM2 that was highlighted by the presence of three alpha-helices encompassing residues 39-47, 49-72, and 80-103, with higher flexibility around the internal Arg(58) site of TM1. RMSD values of individually superimposed helical segments 39-47, 49-72, and 80-103 were 0.25 +/- 0.10 A, 0.40 +/- 0.13 A, and 0.57 +/- 0.19 A, respectively. Several long-range interhelical connectivities supported the folding of TM1-TM2 into a tertiary structure typified by a crossed helix that splays apart toward the extracellular regions and contains considerable flexibility in the G(56)VRSG(60) region. (15)N-relaxation and hydrogen-deuterium exchange data support a stable fold for the TM parts of TM1-TM2, whereas the solvent-exposed segments are more flexible. The NMR structure is consistent with the results of biochemical experiments that identified the ligand-binding site within this region of the receptor.

PubMed: 19383463
DOI: 10.1016/j.bpj.2009.01.012

Experimental method

SOLUTION NMR