PMC:4504005 / 7413-8290 JSONTXT

{"project":"TEST0","denotations":[{"id":"25982113-151-159-8319010","span":{"begin":151,"end":155},"obj":"[\"24115156\", \"24115156\", \"24115156\"]"},{"id":"25982113-234-242-8319011","span":{"begin":292,"end":296},"obj":"[\"21625560\", \"21625560\", \"21625560\"]"},{"id":"25982113-226-234-8319012","span":{"begin":310,"end":314},"obj":"[\"20966082\", \"20966082\", \"20966082\"]"},{"id":"25982113-234-242-8319013","span":{"begin":332,"end":336},"obj":"[\"18801728\", \"18801728\", \"18801728\"]"},{"id":"25982113-226-234-8319014","span":{"begin":338,"end":342},"obj":"[\"19674967\", \"19674967\", \"19674967\"]"},{"id":"25982113-232-240-8319015","span":{"begin":344,"end":348},"obj":"[\"20172855\", \"20172855\", \"20172855\"]"},{"id":"25982113-234-242-8319016","span":{"begin":366,"end":370},"obj":"[\"18375760\", \"18375760\", \"18375760\"]"}],"text":"We previously reported a tethered fusion protein approach to engineer the CLR:RAMP1 and CLR:RAMP2 ECD complexes for crystallization (Moad and Pioszak, 2013), inspired by previous successes using maltose binding protein (MBP) as a “crystallization module” for class B GPCR ECDs (Kumar et al., 2011; Pal et al., 2010; Pioszak et al., 2008, 2009, 2010; Pioszak and Xu, 2008). MBP-RAMP1 or MBP-RAMP2 ECD-CLR ECD fusion proteins in which the two ECDs were covalently tethered with a flexible (Gly-Ser)5 linker were designed to ensure complex stability and enforce 1:1 CLR:RAMP stoichiometry. The tethered RAMP1-CLR ECD fusion was a monomer, whereas the tethered RAMP2-CLR ECD fusion purified as a dimer, but the physiological relevance of oligomerization is unknown. Both proteins selectively bound their respective peptides but failed to yield crystals in the presence of peptides."}

{"project":"2_test","denotations":[{"id":"25982113-24115156-118216112","span":{"begin":151,"end":155},"obj":"24115156"},{"id":"25982113-24115156-118216112","span":{"begin":151,"end":155},"obj":"24115156"},{"id":"25982113-21625560-118216113","span":{"begin":292,"end":296},"obj":"21625560"},{"id":"25982113-21625560-118216113","span":{"begin":292,"end":296},"obj":"21625560"},{"id":"25982113-20966082-118216114","span":{"begin":310,"end":314},"obj":"20966082"},{"id":"25982113-20966082-118216114","span":{"begin":310,"end":314},"obj":"20966082"},{"id":"25982113-18801728-118216115","span":{"begin":332,"end":336},"obj":"18801728"},{"id":"25982113-18801728-118216115","span":{"begin":332,"end":336},"obj":"18801728"},{"id":"25982113-19674967-118216116","span":{"begin":338,"end":342},"obj":"19674967"},{"id":"25982113-19674967-118216116","span":{"begin":338,"end":342},"obj":"19674967"},{"id":"25982113-20172855-118216117","span":{"begin":344,"end":348},"obj":"20172855"},{"id":"25982113-20172855-118216117","span":{"begin":344,"end":348},"obj":"20172855"},{"id":"25982113-18375760-118216118","span":{"begin":366,"end":370},"obj":"18375760"},{"id":"25982113-18375760-118216118","span":{"begin":366,"end":370},"obj":"18375760"},{"id":"25982113-24115156-71076112","span":{"begin":151,"end":155},"obj":"24115156"},{"id":"25982113-24115156-71076112","span":{"begin":151,"end":155},"obj":"24115156"},{"id":"25982113-21625560-71076113","span":{"begin":366,"end":370},"obj":"21625560"},{"id":"25982113-21625560-71076113","span":{"begin":366,"end":370},"obj":"21625560"},{"id":"25982113-20966082-71076113","span":{"begin":366,"end":370},"obj":"20966082"},{"id":"25982113-20966082-71076113","span":{"begin":366,"end":370},"obj":"20966082"},{"id":"25982113-18801728-71076113","span":{"begin":366,"end":370},"obj":"18801728"},{"id":"25982113-18801728-71076113","span":{"begin":366,"end":370},"obj":"18801728"},{"id":"25982113-19674967-71076113","span":{"begin":366,"end":370},"obj":"19674967"},{"id":"25982113-19674967-71076113","span":{"begin":366,"end":370},"obj":"19674967"},{"id":"25982113-20172855-71076113","span":{"begin":366,"end":370},"obj":"20172855"},{"id":"25982113-20172855-71076113","span":{"begin":366,"end":370},"obj":"20172855"},{"id":"25982113-18375760-71076113","span":{"begin":366,"end":370},"obj":"18375760"},{"id":"25982113-18375760-71076113","span":{"begin":366,"end":370},"obj":"18375760"}],"text":"We previously reported a tethered fusion protein approach to engineer the CLR:RAMP1 and CLR:RAMP2 ECD complexes for crystallization (Moad and Pioszak, 2013), inspired by previous successes using maltose binding protein (MBP) as a “crystallization module” for class B GPCR ECDs (Kumar et al., 2011; Pal et al., 2010; Pioszak et al., 2008, 2009, 2010; Pioszak and Xu, 2008). MBP-RAMP1 or MBP-RAMP2 ECD-CLR ECD fusion proteins in which the two ECDs were covalently tethered with a flexible (Gly-Ser)5 linker were designed to ensure complex stability and enforce 1:1 CLR:RAMP stoichiometry. The tethered RAMP1-CLR ECD fusion was a monomer, whereas the tethered RAMP2-CLR ECD fusion purified as a dimer, but the physiological relevance of oligomerization is unknown. Both proteins selectively bound their respective peptides but failed to yield crystals in the presence of peptides."}