Our research objectives are to understand biological processes and phenomena in terms of physical mechanisms and to explore the adaptation of physical techniques to problems in biology. The more specific goals and problems to which we are addressing ourselves are the following: 1.) The mechanism of energy conversion in a protein-chlorophyll complex solubilized frm the membranes of photosynthetic bacteria and the characterization and determination of the structure function relation of this complex called the reaction center (RC).
Specific aims are to: (a) complete the amino acid sequence of the RC protein, (b) to obtain the three-dimensional structure of the RC by X-ray diffraction, (c) to investigate the effect of protein modifications on the structure and funciton of the RC, and (d) to extend the investigation to other membrane proteins. Some of the findings and methodologies are expected to be relevant to other electron-transfer processes, most notably the respiratory chain in mitochondria. 2.) Electronic Structure of Biomolecules: We want to extend our understanding of the non-heme iron center of RCs to the atomic (molecular) level. EPR and ENDOR experiments will be used to determine the parameters characterizing the electronic state of the Fe2+. 3.) Crystallization of Biological Macromolecules, with special emphasis on proteins. Our goal is to understand the detailed mechanisms leading to the formation of single crystals. The three temporal phases of crystallization, i.e., a.) nucleation, b.) post-nucleation growth, and c.) cessation of growth, will be investigated on the water-soluble model compound. lysozyme. We plan to extend the methodology to other proteins, including integral membrane proteins. The long-term objective is to arrive at a systematic procedure to obtain large, high-quality crystals for X-ray diffraction studies.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37GM013191-28
Application #
3484138
Study Section
Special Emphasis Panel (NSS)
Project Start
1976-05-10
Project End
1996-06-30
Budget Start
1992-07-01
Budget End
1993-06-30
Support Year
28
Fiscal Year
1992
Total Cost
Indirect Cost
Name
University of California San Diego
Department
Type
Schools of Arts and Sciences
DUNS #
077758407
City
La Jolla
State
CA
Country
United States
Zip Code
92093
Chirino, A J; Lous, E J; Huber, M et al. (1994) Crystallographic analyses of site-directed mutants of the photosynthetic reaction center from Rhodobacter sphaeroides. Biochemistry 33:4584-93
Paddock, M L; Rongey, S H; McPherson, P H et al. (1994) Pathway of proton transfer in bacterial reaction centers: role of aspartate-L213 in proton transfers associated with reduction of quinoneto dihydroquinone. Biochemistry 33:734-45
McPherson, P H; Schonfeld, M; Paddock, M L et al. (1994) Protonation and free energy changes associated with formation of QBH2 in native and Glu-L212-->Gln mutant reaction centers from Rhodobacter sphaeroides. Biochemistry 33:1181-93
Rongey, S H; Paddock, M L; Feher, G et al. (1993) Pathway of proton transfer in bacterial reaction centers: second-site mutation Asn-M44-->Asp restores electron and proton transfer in reaction centers from the photosynthetically deficient Asp-L213-->Asn mutant of Rhodobacter sphaeroides. Proc Natl Acad Sci U S A 90:1325-9
McPherson, P H; Okamura, M Y; Feher, G (1993) Light-induced proton uptake by photosynthetic reaction centers from Rhodobacter sphaeroides R-26.1. II. Protonation of the state DQAQB2-. Biochim Biophys Acta 1144:309-24
Beroza, P; Fredkin, D R; Okamura, M Y et al. (1991) Protonation of interacting residues in a protein by a Monte Carlo method: application to lysozyme and the photosynthetic reaction center of Rhodobacter sphaeroides. Proc Natl Acad Sci U S A 88:5804-8
Durbin, S D; Feher, G (1990) Studies of crystal growth mechanisms of proteins by electron microscopy. J Mol Biol 212:763-74
Calvo, R; Passeggi, M C; Isaacson, R A et al. (1990) Electron paramagnetic resonance investigation of photosynthetic reaction centers from Rhodobacter sphaeroides R-26 in which Fe2+ was replaced by Cu2+. Determination of hyperfine interactions and exchange and dipole-dipole interactions between Cu2+ and QA- Biophys J 58:149-65
Lubitz, W; Isaacson, R A; Okamura, M Y et al. (1989) ENDOR studies of the intermediate electron acceptor radical anion I-. in Photosystem II reaction centers. Biochim Biophys Acta 977:227-32
Allen, J P; Feher, G; Yeates, T O et al. (1988) Structure of the reaction center from Rhodobacter sphaeroides R-26: protein-cofactor (quinones and Fe2+) interactions. Proc Natl Acad Sci U S A 85:8487-91

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