Summary for 7PVC
| Entry DOI | 10.2210/pdb7pvc/pdb |
| NMR Information | BMRB: 25839 |
| Descriptor | Potassium binding protein Kbp, POTASSIUM ION (2 entities in total) |
| Functional Keywords | potassium ion binding metal ion binding response to potassium ion response to stimulus cytoplasm, metal binding protein |
| Biological source | Escherichia coli (strain K12) |
| Total number of polymer chains | 1 |
| Total formula weight | 17193.33 |
| Authors | Smith, B.O. (deposition date: 2021-10-01, release date: 2021-10-13, Last modification date: 2023-10-11) |
| Primary citation | Torres Caban, C.C.,Yang, M.,Lai, C.,Yang, L.,Subach, F.V.,Smith, B.O.,Piatkevich, K.D.,Boyden, E.S. Tuning the Sensitivity of Genetically Encoded Fluorescent Potassium Indicators through Structure-Guided and Genome Mining Strategies. ACS Sens, 7:1336-1346, 2022 Cited by PubMed Abstract: Genetically encoded potassium indicators lack optimal binding affinity for monitoring intracellular dynamics in mammalian cells. Through structure-guided design and genome mining of potassium binding proteins, we developed green fluorescent potassium indicators with a broad range of binding affinities. KRaION1 (K ratiometric indicator for optical imaging based on mNeonGreen 1), based on the insertion of a potassium binding protein, Kbp, from (Ec-Kbp) into the fluorescent protein mNeonGreen, exhibits an isotonically measured of 69 ± 10 mM (mean ± standard deviation used throughout). We identified Ec-Kbp's binding site using NMR spectroscopy to detect protein-thallium scalar couplings and refined the structure of Ec-Kbp in its potassium-bound state. Guided by this structure, we modified KRaION1, yielding KRaION1/D9N and KRaION2, which exhibit isotonically measured 's of 138 ± 21 and 96 ± 9 mM. We identified four Ec-Kbp homologues as potassium binding proteins, which yielded indicators with isotonically measured binding affinities in the 39-112 mM range. KRaIONs functioned in HeLa cells, but the values differed from the isotonically measured case. We found that, by tuning the experimental conditions, values could be obtained that were consistent and . We thus recommend characterizing potassium indicator in the physiological context of interest before application. PubMed: 35427452DOI: 10.1021/acssensors.1c02201 PDB entries with the same primary citation |
| Experimental method | SOLUTION NMR |
Structure validation
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