Relative binding affinities of chlorophylls in peridinin–chlorophyll–protein reconstituted with heterochlorophyllous mixtures

Abstract

Peridinin-chlorophyll-protein (PCP), containing differently absorbing chlorophyll derivatives, are good models with which to study energy transfer among monomeric chlorophylls (Chls) by both bulk and single-molecule spectroscopy. They can be obtained by reconstituting the N-terminal domain of the protein (N-PCP) with peridinin and chlorophyll mixtures. Upon dimerization of these "half-mers", homo- and heterochlorophyllous complexes are generated, that correspond structurally to monomeric protomers of native PCP from Amphidinium carterae. Heterochlorophyllous complexes contain two different Chls in the two halves of the complete structure. Here, we report reconstitution of N-PCP with binary mixtures of Chl a, Chl b, and [3-acetyl]-Chl a. The ratios of the pigments were varied in the reconstitution mixture, and relative binding constants were determined from quantification of these pigments in the reconstituted PCPs. We find higher affinities for both Chl b and [3-acetyl]-Chl a than for the native pigment, Chl a.

X‐ray structures of the peridinin–chlorophyll‐protein reconstituted with different chlorophylls

“…Polivka et al observed a stochiometric ratio of 1:2 for Chl‐ a :BChl‐ a integration into RFPCP refolded with a 1:1 mixture of the Chls [20]. This observation is confirmed by the biochemical quantification experiments of Brotosudarmo et al who found that acChl‐ a (difference between BChl‐ a and acChl‐ a is the saturation of C7–C8 bond) is integrated twice as often as Chl‐ a [21] . Again, Brotosodarmu et al proposed an H‐bond to account for the observed preference for acChl‐ a over Chl‐ a .…”

“…Brotosudarmo et al have observed a higher binding affinity for Chl‐ b over Chl‐ a in RFPCP in single molecule spectroscopic studies [19] which was confirmed in biochemical quantification experiments [21]. Based on a structural alignment of Chl‐ b onto Chl‐ a , they proposed a hydrogen bond between the formyl group and the C1′‐hydroxyl group of Per‐612 to account for the higher affinity.…”

“…Given this high structural consensus between the Cα atoms it is not surprising that the pigment arrangement is almost identical in all refolded Chl‐RFPCPs (Table 2). Several studies [19–21] have shown that the apparent binding affinities for the Chls with polar substituents at R3 and R7 (Fig. 1D) are higher than for native Chl‐ a .…”

Absorption Enhancement in Peridinin–Chlorophyll–Protein Light-Harvesting Complexes Coupled to Semicontinuous Silver Film

“…The peridinin–chlorophyll–protein complexes used in our experiments were prepared as described [16]. In order to obtain PCP complexes containing Chl, the N-terminal apoprotein was reconstituted with an appropriate mixture of Per and, as desired, Chl or acChl.…”

“…The absorption (and fluorescence excitation) spectrum of the acChl–PCP has an intense, broad band between 400 and 550 nm that is due to Per and two Chl bands at 440 nm (Soret) and 690 nm (Q Y ), the emission spectrum peaks at 690 nm (Fig. 1a) [16].

Fig.

…”