A hereditary spastic paraplegia–associated atlastin variant exhibits defective allosteric coupling in the catalytic core

Abstract

The dynamin-related GTPase atlastin (ATL) catalyzes membrane fusion of the endoplasmic reticulum and thus establishes a network of branched membrane tubules. When ATL function is compromised, the morphology of the endoplasmic reticulum deteriorates, and these defects can result in neurological disorders such as hereditary spastic paraplegia and hereditary sensory neuropathy. ATLs harness the energy of GTP hydrolysis to initiate a series of conformational changes that enable homodimerization and subsequent membrane fusion. Disease-associated amino acid substitutions cluster in regions adjacent to ATL's catalytic site, but the consequences for the GTPase's molecular mechanism are often poorly understood. Here, we elucidate structural and functional defects of an atypical hereditary spastic paraplegia mutant, ATL1-F151S, that is impaired in its nucleotide-hydrolysis cycle but can still adopt a high-affinity homodimer when bound to a transition-state analog. Crystal structures of mutant proteins yielded models of the monomeric pre- and post-hydrolysis states of ATL. Together, these findings define a mechanism for allosteric coupling in which Phe is the central residue in a hydrophobic interaction network connecting the active site to an interdomain interface responsible for nucleotide loading.

Identifying coupled clusters of allostery participants through chemical shift perturbations

“…We expect that multiple NMR lines from allostery participants will likely be observed in many other systems. One recent example has been observed in the GTPase atlastin: Residue F151 displays multiple conformation states in the crystal structures collected in various states and is reasoned to be the connecting residue between the active and nucleotide binding sites (69). Such a "multiple-states" behavior Mutation of T74S causes a strong reduction of allosteric coupling, indicating that the side chain is a crucial player for propagating allosteric interaction and C-type inactivation.…”

The atlastin membrane anchor forms an intramembrane hairpin that does not span the phospholipid bilayer

“…The third class of membrane fusion proteins typified by atlastin and mitofusin are fusion GTPases (2, 5, 6) that use the chemical energy of GTP hydrolysis to drive conformational changes necessary to fuse membranes. Recent work has provided mechanistic insights into the conformational changes that occur during GTP hydrolysis (31,34,(55)(56)(57)(58)), yet the role of the required hydrophobic membrane anchor remains relatively unexplored. Additionally, several proteins have been shown to interact with atlastin, primarily through the hydrophobic membrane anchor, including reticulon/REEP proteins, lunapark, and spastin (6, 7, 17, 25, 37).…”

The structural biology of the dynamin‐related proteins: New insights into a diverse, multitalented family

“…88 In contrast to dynamins, atlastins contain a guanine cap (G5 motif) 89,90 that sequesters the nucleotide from the solvent and utilize a conserved arginine finger in the P-loop (R77) as the charge compensating element ( Figure 7). 91 [94][95][96] Although each is consistent with the respective assays employed, a definitive consensus has yet to be reached in the field.…”

“…These structures suggest a general model for atlastin function in which the extended Form 2 represents a “pre‐fusion” state, with monomers localized in and tethering two distinct membranes, and Forms 1 and 3 represent a “post‐fusion” state, where the conformational rearrangement of the 3HBs facilitates membrane fusion. Different biophysical studies and experimental conditions have yielded alternative models for the timing of these conformational changes relative to atlastin's GTP hydrolysis cycle . Although each is consistent with the respective assays employed, a definitive consensus has yet to be reached in the field.…”