The Reddest Fluorescence of Photosystem I from <i>Halomicronema hongdechloris</i> Comes from Chlorophyll-<i>f</i> Dimer

Chlorophyll-<i>f</i> (Chl-<i>f</i>) is the most red-shifted Chl ever found in nature. It was first found in the cyanobacterium <i>Halomicronema</i> (<i>H.</i>) <i>hongdechloris</i>. This cyanobacterium was found inside rocks called stromatolites. It produces Chl-<i>f</i> to utilize far-red light when grown under far-red light (>700 nm), which is the only light available in a stromatolite. A structural analysis on photosystem I of the <i>H. hongdechloris</i> grown under far-red light showed that among ca. 100 antenna Chls-<i>a</i>, seven are replaced with Chl-<i>f</i>. Three fluorescence peaks at 746, 753, and 810 nm observed at 77 K are attributed to Chl-<i>f</i>. However, it has been unknown which Chl-<i>f</i> is responsible for each red-shifted fluorescence band. Furthermore, the mechanism and pathway of the extreme uphill excitation energy transfer from Chl-<i>f</i> to RC, which was shown to be still at 700 nm, have been a mystery. Here, we employed time-resolved fluorescence spectroscopy, single-PSI fluorescence polarization measurements, and numerical calculations based on the structure of the PSI to assign each far-red emission peak to the Chl-<i>f</i> in the known structure of PSI. Importantly, the reddest emission band at 810 nm was assigned to a dimeric Chl-<i>f</i> composed of f826A and f827A (nomenclature according to Kato <i>Nat. Commun.</i> 2020, 11, 238) located near the RC.