ABSTRACT
Membrane remodeling and repair are essential for all cells. Proteins that perform these functions include Vipp1/IM30 in photosynthetic plastids, PspA in bacteria, and ESCRT-III in eukaryotes. Here, using a combination of evolutionary and structural analyses, we show that these protein families are homologous and share a common ancient evolutionary origin that likely predates the last universal common ancestor. This homology is evident in cryo-electron microscopy structures of Vipp1 rings from the cyanobacterium Nostoc punctiforme presented over a range of symmetries. Each ring is assembled from rungs that stack and progressively tilt to form dome-shaped curvature. Assembly is facilitated by hinges in the Vipp1 monomer, similar to those in ESCRT-III proteins, which allow the formation of flexible polymers. Rings have an inner lumen that is able to bind and deform membranes. Collectively, these data suggest conserved mechanistic principles that underlie Vipp1, PspA, and ESCRT-III-dependent membrane remodeling across all domains of life.
Subject(s)
Bacterial Proteins/metabolism , Cell Membrane/metabolism , Endosomal Sorting Complexes Required for Transport/metabolism , Heat-Shock Proteins/metabolism , Multigene Family , Nostoc/metabolism , Amino Acid Sequence , Animals , Bacterial Proteins/chemistry , Bacterial Proteins/isolation & purification , Bacterial Proteins/ultrastructure , Chickens , Cryoelectron Microscopy , Endosomal Sorting Complexes Required for Transport/chemistry , Evolution, Molecular , Heat-Shock Proteins/chemistry , Heat-Shock Proteins/ultrastructure , Humans , Models, Molecular , Protein Structure, Secondary , Sequence Homology, Amino Acid , ThermodynamicsABSTRACT
Dynamin 1-like proteins (DNM1-L) are mechanochemical GTPases that induce membrane fission in mitochondria and peroxisomes. Their mechanism depends on conformational changes driven by nucleotide and lipid cycling. Here we show the crystal structure of a mitochondrial fission dynamin (CmDnm1) from the algae Cyanidioschyzon merolae. Unlike other eukaryotic dynamin structures, CmDnm1 is in a hinge 1 closed conformation, with the GTPase domain compacted against the stalk. Within the crystal, CmDnm1 packs as a diamond-shaped tetramer that is consistent with an inactive off-membrane state. Crosslinking, photoinduced electron transfer assays, and electron microscopy verify these structures. In vitro, CmDnm1 forms concentration-dependent rings and protein-lipid tubes reminiscent of DNM1-L and classical dynamin with hinge 1 open. Our data provides a mechanism for filament collapse and membrane release that may extend to other dynamin family members. Additionally, hinge 1 closing may represent a key conformational change that contributes to membrane fission.
Subject(s)
Dynamins/metabolism , Mitochondrial Dynamics , Catalysis , Crystallography, X-Ray , Dynamins/chemistry , Molecular Conformation , Reproducibility of ResultsABSTRACT
In bacteria the ability to remodel membrane underpins basic cell processes such as growth, and more sophisticated adaptations like inter-cell crosstalk, organelle specialisation, and pathogenesis. Here, selected examples of membrane remodelling in bacteria are presented and the diverse mechanisms for inducing membrane fission, fusion, and curvature discussed. Compared to eukaryotes, relatively few curvature-inducing proteins have been characterised so far. Whilst it is likely that many such proteins remain to be discovered, it also reflects the importance of alternative membrane remodelling strategies in bacteria where passive mechanisms for generating curvature are utilised.
Subject(s)
Bacteria/ultrastructure , Membranes/ultrastructure , Membrane Proteins/physiology , Membranes/metabolismABSTRACT
Protein phosphorylation is one of the most common post-translational modifications in cell regulatory mechanisms. Dimerization plays also a crucial role in the kinase activity of many kinases, including RAF, CDK2 (cyclin-dependent kinase 2) and EGFR (epidermal growth factor receptor), with heterodimers often being the most active forms. However, the structural and mechanistic details of how phosphorylation affects the activity of homo- and hetero-dimers are largely unknown. Experimentally, synthesizing protein samples with fully specified and homogeneous phosphorylation states remains a challenge for structural biology and biochemical studies. Typically, multiple changes in phosphorylation lead to activation of the same protein, which makes structural determination methods particularly difficult. It is also not well understood how the occurrence of phosphorylation and dimerization processes synergize to affect kinase activities. In the present article, we review available structural data and discuss how MD simulations can be used to model conformational transitions of RAF kinase dimers, in both their phosphorylated and unphosphorylated forms.
