التفاصيل البيبلوغرافية
| العنوان: |
High-affinity tuning of single fluorescent protein-type indicators by flexible linker length optimization in topology mutant. |
| المؤلفون: |
Hara, Yusuke, Ichiraku, Aya, Matsuda, Tomoki, Sakane, Ayuko, Sasaki, Takuya, Nagai, Takeharu, Horikawa, Kazuki |
| المصدر: |
Communications Biology; 6/8/2024, Vol. 7 Issue 1, p1-10, 10p |
| مصطلحات موضوعية: |
FLUORESCENT probes, SECOND messengers (Biochemistry), CELL imaging, TOPOLOGY, FLUORESCENT proteins, INTRAMOLECULAR proton transfer reactions |
| مستخلص: |
Genetically encoded Ca2+ indicators (GECIs) are versatile for live imaging of cellular activities. Besides the brightness and dynamic range of signal change of GECIs, Ca2+ affinity is another critical parameter for successful Ca2+ imaging, as the concentration range of Ca2+ dynamics differs from low nanomolar to sub-millimolar depending on the celltype and organism. However, ultrahigh-affinity GECIs, particularly the single fluorescent protein (1FP)-type, are lacking. Here, we report a simple strategy that increases Ca2+ affinity through the linker length optimization in topology mutants of existing 1FP-type GECIs. The resulting ultrahigh-affinity GECIs, CaMPARI-nano, BGECO-nano, and RCaMP-nano (Kd = 17–25 nM), enable unique biological applications, including the detection of low nanomolar Ca2+ dynamics, highlighting active signaling cells, and multi-functional imaging with other second messengers. The linker length optimization in topology mutants could be applied to other 1FP-type indicators of glutamate and potassium, rendering it a widely applicable technique for modulating indicator affinity. A strategy is developed to obtain ultrahigh-affinity, single fluorescent protein-type calcium indicators (GECIs), based on the optimization of linkers in topology mutants of previously-characterized GECIs. [ABSTRACT FROM AUTHOR] |
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| قاعدة البيانات: |
Complementary Index |
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