Lhcx proteins in centric diatoms

• Applicant: Professorin Dr. Claudia Büchel
• Subject Area: Plant Biochemistry and Biophysics
• Term: from 2014 to 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 256803100

Diatoms are photosynthetic organisms that contribute to about 25% to the yearly biomass production on earth. They possess different proteins in the thylakoid membrane working in light harvesting, called fucoxanthin-chlorophyll-binding proteins (FCP) because of their major pigments. Based on sequence comparison those fall into three groups, Lhcf proteins serving as main light harvesters, Lhcr proteins associated with photosystem I, and Lhcx proteins. Lhcx are involved in protection against a sur-plus of light by increased heat dissipation at the expense of fluorescence emission by a mechanism called non-photochemical quenching (NPQ). The so-called energy dependent qE part of NPQ depends on the transthylakoidal pH gradient, the de-epoxidation of diadinoxanthin to diatoxanthin and on the Lhcx proteins. Recently, we could prove the existence of one of the Lhcx proteins in trimeric FCPa complexes in a centric diatom, and we could show that the fluorescence yield of these complexes depends on the protein-protein distance, the amounts of diatoxanthin bound as well as on the pH. However, diatoms possess more than one Lhcx. Thus we want to study the different Lhcx proteins in the centric diatom Thalassiosira pseudonana, since the genome is sequenced and the alga is transformable. First we want to purify sub-populations of the FCPa complexes from wild type and identify the Lhcx proteins present by mass spectrometry. Having identified those Lhcx present in the different FCPa subpopulations, we want to create transformands of T. pseudonana carrying additional genes with tagged versions of the respective Lhcx proteins. From the transformands the specific FCPa complexes containing these Lhcx should be isolated in a pure state by affinity chromatography, and characterised biochemically. Complexes obtained will then be examined spectroscopically for their fluorescence emission under conditions important for NPQ, like varying protein distances, pH or diatoxanthin content. In addition, knock-down mutants for the same Lhcx will be created and studied for their NPQ capacity by fluorescence kinetics, and their pigment composition will be analysed. Taking together the in vivo and in vitro data we will be able to determine the function of the different Lhcx proteins in NPQ.

• DFG Programme: Research Grants
• Participating Persons: Professorin Dr. Maria Mittag; Professor Dr. Josef Wachtveitl