Park JH, Scheerer P, Hofmann KP, Choe HW, Ernst OP. Crystal structure of the ligand-free G-protein-coupled receptor opsin. Nature. 2008;454(7201):183–7. Epub 2008/06/20. pmid:18563085 The Golden Minigunner is an upgraded version of the Minigunner, obtained from the Golden Skincrate for 50,000 if you own the Minigunner. It can also be bought in the Daily Store for 75,000, being the most expensive item purchasable in the game with coins. All finance and hiring facilities are subject to status and available to over 18s in the UK only (excluding the Channel Islands). Guarantees and indemnities may be required. Finance provided by MINI Financial Services, Summit ONE, Summit Avenue, Farnborough, Hampshire GU14 0FB. Registered Company Number 01288537. Renaud JP, Chari A, Ciferri C, et al. (2018) Cryo-EM in drug discovery: Achievements, limitations and prospects. Nat Rev Drug Discov 17: 471–492. doi: 10.1038/nrd.2018.77

Choe HW, Kim YJ, Park JH, et al. (2011) Crystal structure of metarhodopsin II. Nature 471: 651–655. doi: 10.1038/nature09789 Oldham WM, Hamm HE. Structural basis of function in heterotrimeric G proteins. Q Rev Biophys. 2006;39(2):117–66. pmid:16923326 Kull B, Svenningsson P, Fredholm BB. Adenosine A(2A) receptors are colocalized with and activate g(olf) in rat striatum. Molecular pharmacology. 2000;58(4):771–7. Epub 2000/09/22. pmid:10999947 The values shown for fuel consumption and CO2 emissions are calculated in accordance with the measurement method prescribed in the European Regulation (EC) 715/2007 in the version applicable at the time of approval.

Rasmussen SG, Devree BT, Zou Y, et al. (2011) Crystal structure of the beta2 adrenergic receptor-Gs protein complex. Nature 477: 549–555. doi: 10.1038/nature10361

As previously mentioned,you really need an RTA or RDA to get the best out of the SX Mini G Class V2, especially dual coil round wire tanks, I used the OXVA Arbiter 2 which is quite large but even that looked small on the mod.You can comfortably go up to a 30mm tank, but I found anything less than 26mm started to look a little lost on it, especially a 24mm RDA I used for test TC mode. The G-zone is simply an area of tissue that some people find pleasurable, while others don’t,” Corrado says. “That pleasure has more to do with the type of pressure that is being put on the clitoral bulbs and the Skene’s glands, rather than an entirely separate structure.” The Skene’s glands are two glands located on either side of the urethra (a tube that connects to the bladder for urination) which are thought to be responsible for vaginal secretions during arousal. Archetypical members from each Gα family were selected and include the following: G olf from the G s family, G i1, G o1, G z and G t from the G i family, G q and G 16 from the G q/11 family, and G 12 from the G 12/13 family. The mutations required to convert Gα s into mini-G s were transferred en bloc to the selected Gα proteins to produce a mini-G protein version of each ( Fig 2). These mutations were the following: (i) deletion of all amino acid residues N-terminal of Ile/Leu HN43; (ii) deletion of the α-helical domain between residues H H1S2.12 and the Thr, three residues N-terminal to Ile S2.1, and replacement with an 8 amino acid residue linker; (iii) deletion of 10 amino acid residues of switch III between Tyr S4H3.4 and Asn/Ser S4H3.15; (iv) mutating 7 residues to D49 S1H1.3, N50 S1H1.4, D249 S4.7, D252 S4H3.3, D272 H3.8, A372 H5.4, I375 H5.7. Residue numbers are for Gα s and superscripts refer to the CGN system for comparing residues in G proteins [ 6]. Initial characterization of each mini-G protein was performed by assessing expression in Escherichia coli and purification by Ni 2+-affinity chromatography and size exclusion chromatography (SEC). Four out of the eight engineered mini-G proteins (mini-G olf, mini-G i1, mini-G o1 and mini-G 12) fulfilled these initial criteria i. e. they were all stable enough in their basal conformation to allow high-yield expression and purification. The yield of purified mini-G protein per litre of culture and their stability as measured by differential scanning fluorimetry (in parentheses) are as follows: mini-G s, 100 mg/L (65°C); mini-G olf, 80 mg/L (65°C); mini-G o1 100 mg/L (64°C); mini-G 12 25 mg/L (73°C). The worst expressed of the four new mini-G proteins was mini-G i1, so an additional mutation G217D was incorporated and the truncation at the N-terminus shortened, which increased the yield of pure protein to 12 mg/L, although the stability was only 48°C. Thus, mini-G olf, mini G i1, mini-G o1 and mini-G 12 were all of sufficient stability to be used to test their ability to couple to relevant GPCRs. The amino acid sequences of the mini-G proteins are given in S1 Fig. Zhang X, Stevens RC, Xu F. The importance of ligands for G protein-coupled receptor stability. Trends in biochemical sciences. 2015;40(2):79–87. pmid:25601764 Landau EM, Rosenbusch JP (1996) Lipidic cubic phases: A novel concept for the crystallization of membrane proteins. Proc Natl Acad Sci USA 93: 14532–14535. doi: 10.1073/pnas.93.25.14532

Results and discussion

Warne T, Serrano-Vega MJ, Baker JG, Moukhametzianov R, Edwards PC, Henderson R, et al. Structure of a beta1-adrenergic G-protein-coupled receptor. Nature. 2008;454(7203):486–91. Epub 2008/07/03. pmid:18594507 Green SA, Holt BD, Liggett SB (1992) Beta 1- and beta 2-adrenergic receptors display subtype-selective coupling to Gs. Mol Pharmacol 41: 889–893.