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1. Design of a light and Ca ²⁺ switchable organic–peptide hybrid

   https://doi.org/10.1073/pnas.2411316122  

   Khaleel, Zinah_Hilal; No, Young_Hyun; Kim, Nam_Hyeong; Bae, Do_Hyun; Wu, Yibing; Kim, Suhyeon; Choi, Hojae; Lee, Da_Eun; Jeong, Se_Yun; Ko, Yoon-Joo; et al (January 2025, Proceedings of the National Academy of Sciences)

   The design of organic–peptide hybrids has the potential to combine our vast knowledge of protein design with small molecule engineering to create hybrid structures with complex functions. Here, we describe the computational design of a photoswitchable Ca²⁺-binding organic–peptide hybrid. The designed molecule, designated Ca²⁺-binding switch (CaBS), combines an EF-hand motif from classical Ca²⁺-binding proteins such as calmodulin with a photoswitchable group that can be reversibly isomerized between a spiropyran (SP) and merocyanine (MC) state in response to different wavelengths of light. The MC/SP group acts both as a photoswitch as well as an optical sensor of Ca²⁺binding. Photoconversion of the SP to the corresponding MC unmasks an acidic phenol, which CaBS uses as an integral part of both its Ca²⁺-binding site as well as its tertiary and quaternary structure. By design, the SP state of CaBS is monomeric, while the Ca²⁺-bound form of the MC state is an obligate dimer, with two Ca²⁺-binding sites formed at the interface of a domain-swapped dimer. Thus, light and Ca²⁺were expected to serve as an “AND gate” that powers a change in backbone structure/dynamics, oligomerization state, and fluorescence properties of the designed molecule. CaBS was designed using Rosetta and molecular dynamics simulations, and experimentally characterized by nuclear magnetic resonance, isothermal titration calorimetry, and optical titrations. These data illustrate the potential of combining small molecule engineering with de novo protein design to develop sensors whose conformation, association state, and optical properties respond to multiple environmental cues. 

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2. The role of Ca ²⁺ and protein scaffolding in the formation of nature’s water oxidizing complex

   https://doi.org/10.1073/pnas.2011315117  

   Avramov, Anton P.; Hwang, Hong J.; Burnap, Robert L. (November 2020, Proceedings of the National Academy of Sciences)

   null (Ed.)

   Photosynthetic O 2 evolution is catalyzed by the Mn 4 CaO 5 cluster of the water oxidation complex of the photosystem II (PSII) complex. The photooxidative self-assembly of the Mn 4 CaO 5 cluster, termed photoactivation, utilizes the same highly oxidizing species that drive the water oxidation in order to drive the incorporation of Mn 2+ into the high-valence Mn 4 CaO 5 cluster. This multistep process proceeds with low quantum efficiency, involves a molecular rearrangement between light-activated steps, and is prone to photoinactivation and misassembly. A sensitive polarographic technique was used to track the assembly process under flash illumination as a function of the constituent Mn 2+ and Ca 2+ ions in genetically engineered membranes of the cyanobacterium Synechocystis sp. PCC6803 to elucidate the action of Ca 2+ and peripheral proteins. We show that the protein scaffolding organizing this process is allosterically modulated by the assembly protein Psb27, which together with Ca 2+ stabilizes the intermediates of photoactivation, a feature especially evident at long intervals between photoactivating flashes. The results indicate three critical metal-binding sites: two Mn and one Ca, with occupation of the Ca site by Ca 2+ critical for the suppression of photoinactivation. The long-observed competition between Mn 2+ and Ca 2+ occurs at the second Mn site, and its occupation by competing Ca 2+ slows the rearrangement. The relatively low overall quantum efficiency of photoactivation is explained by the requirement of correct occupancy of these metal-binding sites coupled to a slow restructuring of the protein ligation environment, which are jointly necessary for the photooxidative trapping of the first stable assembly intermediate. 

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3. Activity-dependent endoplasmic reticulum Ca ²⁺ uptake depends on Kv2.1-mediated endoplasmic reticulum/plasma membrane junctions to promote synaptic transmission

   https://doi.org/10.1073/pnas.2117135119  

   Panzera, Lauren C.; Johnson, Ben; Quinn, Josiah A.; Cho, In Ha; Tamkun, Michael M.; Hoppa, Michael B. (July 2022, Proceedings of the National Academy of Sciences)

   The endoplasmic reticulum (ER) forms a continuous and dynamic network throughout a neuron, extending from dendrites to axon terminals, and axonal ER dysfunction is implicated in several neurological disorders. In addition, tight junctions between the ER and plasma membrane (PM) are formed by several molecules including Kv2 channels, but the cellular functions of many ER-PM junctions remain unknown. Recently, dynamic Ca 2+ uptake into the ER during electrical activity was shown to play an essential role in synaptic transmission. Our experiments demonstrate that Kv2.1 channels are necessary for enabling ER Ca 2+ uptake during electrical activity, as knockdown (KD) of Kv2.1 rendered both the somatic and axonal ER unable to accumulate Ca 2+ during electrical stimulation. Moreover, our experiments demonstrate that the loss of Kv2.1 in the axon impairs synaptic vesicle fusion during stimulation via a mechanism unrelated to voltage. Thus, our data demonstrate that a nonconducting role of Kv2.1 exists through its binding to the ER protein VAMP-associated protein (VAP), which couples ER Ca 2+ uptake with electrical activity. Our results further suggest that Kv2.1 has a critical function in neuronal cell biology for Ca 2+ handling independent of voltage and reveals a critical pathway for maintaining ER lumen Ca 2+ levels and efficient neurotransmitter release. Taken together, these findings reveal an essential nonclassical role for both Kv2.1 and the ER-PM junctions in synaptic transmission. 

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4. Plant “helper” immune receptors are Ca ²⁺ -permeable nonselective cation channels

   https://doi.org/10.1126/science.abg7917  

   Jacob, Pierre; Kim, Nak Hyun; Wu, Feihua; El-Kasmi, Farid; Chi, Yuan; Walton, William G.; Furzer, Oliver J.; Lietzan, Adam D.; Sunil, Sruthi; Kempthorn, Korina; et al (June 2021, Science)

   Plant nucleotide-binding leucine-rich repeat receptors (NLRs) regulate immunity and cell death. InArabidopsis, a subfamily of “helper” NLRs is required by many “sensor” NLRs. Active NRG1.1 oligomerized, was enriched in plasma membrane puncta, and conferred cytoplasmic calcium ion (Ca²⁺) influx in plant and human cells. NRG1.1-dependent Ca²⁺influx and cell death were sensitive to Ca²⁺channel blockers and were suppressed by mutations affecting oligomerization or plasma membrane enrichment. Ca²⁺influx and cell death mediated by NRG1.1 and ACTIVATED DISEASE RESISTANCE 1 (ADR1), another helper NLR, required conserved negatively charged N-terminal residues. Whole-cell voltage-clamp recordings demonstrated thatArabidopsishelper NLRs form Ca²⁺-permeable cation channels to directly regulate cytoplasmic Ca²⁺levels and consequent cell death. Thus, helper NLRs transduce cell death signals directly. 

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5. Resource limitation determines realized thermal performance of consumers in trophodynamic models

   https://doi.org/10.1111/ele.14086  

   Vinton, Anna C.; Vasseur, David A. (August 2022, Ecology Letters)

   Abstract Recent work has demonstrated that changes in resource availability can alter a consumer's thermal performance curve (TPC). When resources decline, the optimal temperature and breadth of thermal performance also decline, leading to a greater risk of warming than predicted by static TPCs. We investigate the effect of temperature on coupled consumer‐resource dynamics, focusing on the potential for changes in the consumer TPC to alter extinction risk. Coupling consumer and resource dynamics generally reduces the potential for resource decline to exacerbate the effects of warming via changes to the TPC due to a reduction in top‐down control when consumers near the limits of their thermal performance curve. However, if resources are more sensitive to warming, consumer TPCs can be reshaped by declining resources, leading to increased extinction risk. Our work elucidates the role of top‐down and bottom‐up regulation in determining the extent to which changes in resource density alter consumer TPCs. 

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