Ion conductivity and the gating characteristics of tetrameric glutamate receptor ion channels are determined by their subunit composition. Competitive homo- and hetero-dimerization of their amino-terminal domains (ATDs) is a key step controlling assembly. Here we measured systematically the thermodynamic stabilities of homodimers and heterodimers of kainate and AMPA receptors using fluorescence-detected sedimentation velocity analytical ultracentrifugation. Measured affinities span many orders of magnitude, and complexes show large differences in kinetic stabilities. The association of kainate receptor ATD dimers is generally weaker than the association of AMPA receptor ATD dimers, but both show a general pattern of increased heterodimer stability as compared to the homodimers of their constituents, matching well physiologically observed receptor combinations. The free energy maps of AMPA and kainate receptor ATD dimers provide a framework for the interpretation of observed receptor subtype combinations and possible assembly pathways.
- Peter Schuck
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Gary L Westbrook, Vollum Institute, United States
This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.
Carotenoid (Car) pigments perform central roles in photosynthesis-related light harvesting (LH), photoprotection, and assembly of functional pigment-protein complexes. However, the relationships between Car depletion in the LH, assembly of the prokaryotic reaction center (RC)-LH complex, and quinone exchange are not fully understood. Here, we analyzed native RC-LH (nRC-LH) and Car-depleted RC-LH (dRC-LH) complexes in Roseiflexus castenholzii, a chlorosome-less filamentous anoxygenic phototroph that forms the deepest branch of photosynthetic bacteria. Newly identified exterior Cars functioned with the bacteriochlorophyll B800 to block the proposed quinone channel between LHαβ subunits in the nRC-LH, forming a sealed LH ring that was disrupted by transmembrane helices from cytochrome c and subunit X to allow quinone shuttling. dRC-LH lacked subunit X, leading to an exposed LH ring with a larger opening, which together accelerated the quinone exchange rate. We also assigned amino acid sequences of subunit X and two hypothetical proteins Y and Z that functioned in forming the quinone channel and stabilizing the RC-LH interactions. This study reveals the structural basis by which Cars assembly regulates the architecture and quinone exchange of bacterial RC-LH complexes. These findings mark an important step forward in understanding the evolution and diversity of prokaryotic photosynthetic apparatus.
Ca2+/calmodulin-dependent protein kinase II (CaMKII) is essential for long-term potentiation (LTP) of excitatory synapses that is linked to learning and memory. In this study, we focused on understanding how interactions between CaMKIIα and the actin-crosslinking protein α-actinin-2 underlie long-lasting changes in dendritic spine architecture. We found that association of the two proteins was unexpectedly elevated within 2 minutes of NMDA receptor stimulation that triggers structural LTP in primary hippocampal neurons. Furthermore, disruption of interactions between the two proteins prevented the accumulation of enlarged mushroom-type dendritic spines following NMDA receptor activation. α-Actinin-2 binds to the regulatory segment of CaMKII. Calorimetry experiments, and a crystal structure of α-actinin-2 EF hands 3 and 4 in complex with the CaMKII regulatory segment, indicate that the regulatory segment of autoinhibited CaMKII is not fully accessible to α-actinin-2. Pull-down experiments show that occupation of the CaMKII substrate-binding groove by GluN2B markedly increases α-actinin-2 access to the CaMKII regulatory segment. Furthermore, in situ labelling experiments are consistent with the notion that recruitment of CaMKII to NMDA receptors contributes to elevated interactions between the kinase and α-actinin-2 during structural LTP. Overall, our study provides new mechanistic insight into the molecular basis of structural LTP and reveals an added layer of sophistication to the function of CaMKII.