The Loroxanthin Cycle: A New Type of Xanthophyll Cycle in Green Algae (Chlorophyta).

Xanthophyll cycles (XC) have proven to be major contributors to photoacclimation for many organisms. This work describes a light-driven XC operating in the chlorophyte Chlamydomonas reinhardtii and involving the xanthophylls Lutein (L) and Loroxanthin (Lo). Pigments were quantified during a switch from high to low light (LL) and at different time points from cells grown in Day/Night cycle. Trimeric LHCII was purified from cells acclimated to high or LL and their pigment content and spectroscopic properties were characterized. The Lo/(L + Lo) ratio in the cells varies by a factor of 10 between cells grown in low or high light (HL) leading to a change in the Lo/(L + Lo) ratio in trimeric LHCII from .5 in low light to .07 in HL. Trimeric LhcbMs binding Loroxanthin have 5 ± 1% higher excitation energy (EE) transfer (EET) from carotenoid to Chlorophyll as well as higher thermo- and photostability than trimeric LhcbMs that only bind Lutein. The Loroxanthin cycle operates on long time scales (hours to days) and likely evolved as a shade adaptation. It has many similarities with the Lutein-epoxide - Lutein cycle (LLx) of plants.

PSI of the Colonial Alga Botryococcus braunii Has an Unusually Large Antenna Size.

PSI is an essential component of the photosynthetic apparatus of oxygenic photosynthesis. While most of its subunits are conserved, recent data have shown that the arrangement of the light-harvesting complexes I (LHCIs) differs substantially in different organisms. Here we studied the PSI-LHCI supercomplex of Botryococccus braunii, a colonial green alga with potential for lipid and sugar production, using functional analysis and single-particle electron microscopy of the isolated PSI-LHCI supercomplexes complemented by time-resolved fluorescence spectroscopy in vivo. We established that the largest purified PSI-LHCI supercomplex contains 10 LHCIs (∼240 chlorophylls). However, electron microscopy showed heterogeneity in the particles and a total of 13 unique binding sites for the LHCIs around the PSI core. Time-resolved fluorescence spectroscopy indicated that the PSI antenna size in vivo is even larger than that of the purified complex. Based on the comparison of the known PSI structures, we propose that PSI in B. braunii can bind LHCIs at all known positions surrounding the core. This organization maximizes the antenna size while maintaining fast excitation energy transfer, and thus high trapping efficiency, within the complex.

Light Acclimation of the Colonial Green Alga Botryococcus braunii Strain Showa.

In contrast to single cellular species, detailed information is lacking on the processes of photosynthetic acclimation for colonial algae, although these algae are important for biofuel production, ecosystem biodiversity, and wastewater treatment. To investigate differences between single cellular and colonial species, we studied the regulation of photosynthesis and photoprotection during photoacclimation for the colonial green alga Botryococcus braunii and made a comparison with the properties of the single cellular species Chlamydomonas reinhardtii We show that B. braunii shares some high-light (HL) photoacclimation strategies with C. reinhardtii and other frequently studied green algae: decreased chlorophyll content, increased free carotenoid content, and increased nonphotochemical quenching (NPQ). Additionally, B. braunii has unique HL photoacclimation strategies, related to its colonial form: strong internal shading by an increase of the colony size and the accumulation of extracellular echinenone (a ketocarotenoid). HL colonies are larger and more spatially heterogenous than low-light colonies. Compared with surface cells, cells deeper inside the colony have increased pigmentation and larger photosystem II antenna size. The core of the largest of the HL colonies does not contain living cells. In contrast with C. reinhardtii, but similar to other biofilm-forming algae, NPQ capacity is substantial in low light. In HL, NPQ amplitude increases, but kinetics are unchanged. We discuss possible causes of the different acclimation responses of C. reinhardtii and B. braunii Knowledge of the specific photoacclimation processes for this colonial green alga further extends the view of the diversity of photoacclimation strategies in photosynthetic organisms.

Light-harvesting complexes of Botryococcus braunii.

The colonial green alga Botryococcus braunii (BB) is a potential source of biofuel due to its natural high hydrocarbon content. Unfortunately, its slow growth limits its biotechnological potential. Understanding its photosynthetic machinery could help to identify possible growth limitations. Here, we present the first study on BB light-harvesting complexes (LHCs). We purified two LHC fractions containing the complexes in monomeric and trimeric form. Both fractions contained at least two proteins with molecular weight (MW) around 25 kDa. The chlorophyll composition is similar to that of the LHCII of plants; in contrast, the main xanthophyll is loroxanthin, which substitutes lutein in most binding sites. Circular dichroism and 77 K absorption spectra lack typical differences between monomeric and trimeric complexes, suggesting that intermonomer interactions do not play a role in BB LHCs. This is in agreement with the low stability of the BB LHCII trimers as compared to the complexes of plants, which could be related to loroxanthin binding in the central (L1 and L2) binding sites. The properties of BB LHCII are similar to those of plant LHCII, indicating a similar pigment organization. Differences are a higher content of red chlorophyll a, similar to plant Lhcb3. These differences and the different Xan composition had no effect on excitation energy transfer or fluorescence lifetimes, which were similar to plant LHCII.

Functionalization of oxidases with peroxidase activity creates oxiperoxidases: a new breed of hybrid enzyme capable of cascade chemistry.

The covalent flavoprotein alditol oxidase (AldO) from Streptomyces coelicolor A3(2) was endowed with an extra catalytic functionality by fusing it to a microperoxidase. Purification of the construct resulted in the isolation of a synthetic bifunctional enzyme that was both fully covalently flavinylated and heminylated: an oxiperoxidase. Characterization revealed that both oxidase and peroxidase functionalities were active, with the construct functioning as a single-component xylitol biosensor. In an attempt to reduce the size of the oxidase-peroxidase fusion, we replaced portions of the native AldO sequence with the bacterial cytochrome c CXXCH heme-binding motif. By mutating only three residues of the AldO protein we were able to create a functional oxidase-peroxidase hybrid.