This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.A central goal of our program is to understand how protein structure controls long-range electron transfer (ET) reactions. Three systems were studied this past year - 1) Interprotein ET in complexes of cytochrome c (Cc) with cytochrome c peroxidase (CcP) and 2) Inter- and intra protein ET reactions in metal-modified azurins (Az) 3) Inter-cofactor ET in prolyl 4-hydroxylase (P4H).1)CcP:Cc - We determined structures of Zn-porphyrin containing CcP with Cc from yeast (yCc) and Cc from horse heart (hCc). YCc and hCc have different modes of interaction with CcP. Electron transfer rates measured directly in these crystals confirm for the first time that the crystal structures represent associations that govern solution reactivity. Moreover ET rate calculations based these new structures indicate that the observed reactivity can only be rationalized if conformational change gates ET. We have also collected diffraction data on complexes that contain site-directed mutants of Cc that drastically alter electron transfer reactivity.2) Az - We determined the high resolution structure of a Re-modified azurin in which a very stable tryptophan radical forms by photochemically triggered oxidative electron transfer to the Re. Surprisingly, the Trp site of the stable radical is solvent exposed and the Trp indole is found in multiple conformations. We have also determined an atomic resolution structure (0.98 resolution) of a Cu and Zn-containing azurin. These are the first atomic resolution structures of a blue-copper protein.3) P4H - We have progressed towards determining the first structure for a collegen-type P4H, an enzyme where inter-cofactor ET rescues oxidatively trapped states. We have now collected diffraction data on three different crystal forms and have improved our native data to 2.4 resolution. We have collected anomalous scattering data from Pb and W derivatives and have an interpretable electron density map. Structure determination is underway.
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