Characterization of mutations at the Glutamate 339 residue in the Photosystem II protein CP 43 in Synechocystis / by Ethan Dunn.
| Author/creator | Dunn, Ethan author. |
| Other author | Putnam-Evans, Cindy, degree supervisor. |
| Other author | East Carolina University. Department of Biology. |
| Format | Theses and dissertations |
| Production | 2008. |
| Description | 83 leaves : illustrations (some color) ; 29 cm |
| Supplemental Content | Access via ScholarShip |
| Subjects |
| Summary | Photosynthesis is the process that cyanobacteria, algae, and higher plants use to convert light energy into chemical energy in the form of carbohydrates. Photosystem II (PS H) is a multisubunit protein-cofactor complex embedded in the thylakoid membrane that catalyzes the light dependent oxidation of water and reduction of plastoquinone with the concomitant evolution of molecular oxygen. One component of PS H is the psbC gene product CP 43. CP 43 is a chlorophyll a binding core antenna protein that is essential for oxygen evolution and the proper assembly and function of PS H. CP 43 has six transmembrane helices connected by five hydrophilic loop regions, the largest being large extrinsic loop E, which extends into the thylakoid lumen. The thylakoid lumen is the site of water oxidation and loop E is proposed to provide direct ligands to the manganese cluster in the oxygen-evolving complex (OEC). The 3.5 angstrom crystal structure of PS II (Ferreira et al., 2004) proposes Glu339 to be a direct bidentate ligand to one Mn atom of the cluster; the most recent crystal structure by Loll et al., (2005) proposes Glu339 to ligate both the 2nd and 3rd Mn atoms. These structures are likely inaccurate due to damage to the Mn cluster during x-ray exposure (Loll et al, 2005 and Pushkar et al., 2006). In this current study, four site-directed mutations at Glu339 (E339A, E339D, E339H, and E339R) were characterized for growth, oxygen-evolving activity, photoinactivation, and variable fluorescence. Protein content of PS 11 mutant particles was examined by LDS-PAGE and immunoblotting. Interestingly, an E339D mutant exhibited a near PS 11 minus phenotype while an E339R mutant exhibited near-normal PS II function. Another mutant, E339H, showed a decrease in PS II function but was still somewhat functional and an E339A mutant exhibited a complete loss of growth and oxygen-evolving ability. A fifth mutant, E339Q, the only Glu339 mutant to be studied to date by Rosenberg et al, (1999), was also characterized for comparison and as a secondary control; this mutant exhibited a severe decrease in PS 11 function. All mutants assembled significant numbers of PSII centers in their membranes. The data obtained here support a role for Glu339 in Mn ligation, however, further studies need to be conducted to assign an exact role for Glu339 in the water splitting mechanism. |
| General note | Presented to the faculty of the Department of Biology. |
| General note | Advisor: Cindy Putnam-Evans |
| Dissertation note | M.S. East Carolina University 2008 |
| Bibliography note | Includes bibliographical references (leaves 77-83). |
| Genre/form | Academic theses. |
| Genre/form | Academic theses. |
| Genre/form | Thèses et écrits académiques. |