Protein and lipid dynamics in photosynthetic thylakoid membranes probed by in-situ solid-state NMR

F. Azadi Chegenia, K.B. Sai Sankar Guptaa, G. Perinb, D. Simionatob, T. Morosinottob, A. Pandita
aLeiden Institute of Chemistry, Solid State NMR, Leiden, Netherlands; bUniversity of Padova, Department of Biology, Padova, Italy

In oxygenic photosynthesis, sophisticated regulation mechanisms have evolved to enable the splitting of water via P680, the strongest oxidizer found in Nature, while preventing the system from photo damage. Under excess light, remodeling of the photosynthetic membrane takes place and molecular switching of light-harvesting antenna proteins into a photoprotective, light-quenching state. Photosynthetic thylakoid membranes are densely packed with proteins (about 70%) and are abundant with proteins from the light-harvesting multigene family.

Here, we explore the use of 13C solid-state NMR on whole thylakoid membranes to gain insight in protein and lipid molecular dynamics in situ. We analyzed Chlamydomonas reinhardtii (Cr.) thylakoid membranes from wild type (WT) and npq2 mutants that accumulate zeaxanthin, testing the ability of 13C NMR spectroscopy for screening molecular membrane characteristics of WT cells and mutants with altered photo protective properties. To separate and quantify rigid and dynamic membrane components, cross polarization, through bond and direct polarization transfer experiments were performed at different temperatures. In addition, T relaxation experiments were performed to determine the backbone dynamics of protein complexes inside the membranes. Results show that we can separate protein and lipid components of our membrane samples by their differential molecular dynamics. A comparison of WT and npq2 mutant membranes, show that the npq2 mutants have increased lipid, but decreased protein dynamics of their thylakoid membranes.