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The late stages of Golgi maturation involve a series of sequential trafficking events in which cargo-laden vesicles are produced and targeted to multiple distinct subcellular destinations. Each of these vesicle biogenesis events requires activation of an Arf GTPase by the Sec7/BIG guanine nucleotide exchange factor (GEF). Sec7 localization and activity is regulated by autoinhibition, positive feedback, and interaction with other GTPases. Although these mechanisms have been characterized biochemically, we lack a clear picture of how GEF localization and activity is modulated by these signals. Here, we report the cryogenic electron microscopy structure of full-length Sec7 in its autoinhibited form, revealing the architecture of its multiple regulatory domains. We use functional experiments to determine the basis for autoinhibition and use structural predictions to produce a model for an active conformation of the GEF that is supported empirically. This study therefore elucidates the conformational transition that Sec7 undergoes to become active on the organelle membrane surface. 

Kohinoor is a positive reversibly switching fluorescent protein. Ultrafast electronic and vibrational spectroscopy suggest that the chromophore switching mechanism is steered by dynamics in surrounding protein residues.