Dynamics in natural photosynthetic biomaterials [electronic resource]

Photosynthesis, a biological process essential to most life on Earth, converts light energy to chemical energy, generates fixed carbon in the form polysaccharides and lipids, and oxidizes water to yield oxygen, protons and electrons. The light-dependent reactions of photosynthesis reside in pigment-protein complexes of the photosynthetic or thylakoid membranes where their activities are integrated in a dynamic manner that facilitates tailoring of photosynthetic activities to specific environmental conditions. The environmental changes could be rapid such as fluctuations in light intensities experienced by photosynthetic organisms because of the shading associated with passing clouds. More gradual changes in light intensity, temperature and humidity occur over the diel cycle. Several dynamic modifications in the composition and arrangement of integral photosynthetic complexes enable plants and algae to cope with changing light conditions. However, these dynamic changes have not been observed in real time at the nanoscale level. This dissertation describes an approach to examine the dynamics of photosynthetic pigment-protein complexes in plants and green algae. Atomic Force Microscopy (AFM) in an aqueous medium was used to visualize these complexes at high resolution in their native thylakoid membranes. For plants (spinach), liquid AFM was used in contact mode to examine the structure of grana membranes, which contain stacked portions of thylakoid membranes enriched in photosystem II (PSII). This work provides the first high resolution surface topography of lumen-protruding domains of PSII reaction centers, and allows inner antenna chlorophyll binding proteins to be distinguished from the extrinsic subunits of PSII that comprise the oxygen evolving complex (PSII-OEC). Furthermore, a novel particle that protrudes on the stromal side of thylakoid membrane was identified as the stroma-exposed domain of photosystem I (PSI). Liquid AFM was also used in tapping or intermittent-contact mode to examine the structure of photosynthetic complexes in algal thylakoid membranes. The unicellular green alga, Chlamydomonas reinhardtii (Chlamydomonas) was used for these studies because it is a powerful biochemical and genetic system for dissecting photosynthesis, with a fully sequenced genome and many photosynthetically-deficient strains. Using a thylakoid isolation procedure that I specifically developed, the organization of photosynthetic complexes on Chlamydomonas thylakoids was, for the first time, characterized using AFM imaging. Examination of various mutants of this alga coupled to the high-resolution imaging capability of liquid AFM, allowed us to distinguish the lumen-protruding domains of the cytochrome b6f (Cyt b6f) from those of PSII (PSII-OEC); Cyt b6f and PSII coexist in PSII-rich regions of Chlamydomonas thylakoid membranes. This information was used to analyze AFM images obtained in studies of state transition, a major photoacclimation process that occurs in many photosynthetic organisms. Although unable to observe the real time dynamics, we were able to detail the spatial reorganization of photosynthetic complexes on Chlamydomonas thylakoid membranes upon state transitions. With our ability to generate high spatial resolution images at near physiological conditions using liquid AFM, we can examine various physiologically processes in addition to photosynthesis (e.g. respiration, metabolite transport, ribosome function) in various biological systems, although we would focus on those systems with well characterized genetics and full genome sequences.

Keywords: Materials Science and Engineering.
Advisor: Grossman Arthur (Arthur R.) primary advisor. Thesis advisor
Prinz F B.
Boxer Steven G. (Steven George)
Melosh Nicholas A. advisor. Thesis advisor