Abstract
GRASPs are proteins involved in cell processes that seem paradoxical: responsible for shaping the Golgi cisternae and involved in unconventional secretion mechanisms that bypass the Golgi. Despite its physiological relevance, there is still a considerable lack of studies on full-length GRASPs. Our group has previously reported an unexpected behavior of the full-length GRASP from the fungus C. neoformans: its intrinsically-disordered characteristic. Here, we generalize this finding by showing that it is also observed in the GRASP from S. cerevisae (Grh1), which strongly suggests it might be a general property within the GRASP family. Furthermore, Grh1 is also able to form amyloid-like fibrils either upon heating or when submitted to changes in the dielectric constant of its surroundings, a condition that is experienced by the protein when in close contact with membranes of cell compartments, such as the Golgi apparatus. Intrinsic disorder and fibril formation can thus be two structural properties exploited by GRASP during its functional cycle.
Highlights
The Golgi complex is composed of a series of cisternal membranes opposed to one another to form stacks[1]
We investigated the biophysical and biochemical features of Grh[1] and the isolated GRASP domain by circular dichroism (CD), fluorescence and optical spectroscopies, differential scanning calorimetry (DSC), computational predictions and established that Grh[1] is a molten globule-like protein, making it a member of the collapsed intrinsically disordered protein (IDP) family
We have described a biophysical characterization of Grh[1] and its GRASP domain that revealed two significant aspects about Grh[1], which are most likely linked: the presence of multiple intrinsically disordered regions that confers to Grh[1] a molten globule-like feature and the capability of forming amyloid fibrils upon mild denaturing conditions, in an SPR-independent fashion
Summary
The Golgi complex is composed of a series of cisternal membranes opposed to one another to form stacks[1]. Details of the involvement of GRASPs in membrane trafficking and other functions in mammalian cells have been reported by researchers using model organisms, such as the yeast Saccharomyces cerevisiae. Saccharomyces cerevisiae has the basic organization of its Golgi cisternae, only 40% of the cisternae are in stacks and the stacks are never found linked to each other[5] This budding yeast contains a single GRASP65 homolog, known as Grh[1], which localizes in compartments of the early secretory pathway[14]. IDPs are proteins involved in a large set of functions and characterized by regions of high polypeptide mobility, and without a well-defined 3D structure[16,17]. The structural flexibility of IDPs allows a broad functional repertoire and a number of interaction partners[19] to act and to influence protein function in different processes, such as transcriptional regulation, translation, cellular signal transduction, and storage of small molecules[20]
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