2yao

Publication Abstract from PubMed

During X-ray data collection from a multicopper oxidase (MCO) crystal, electrons and protons are mainly released into the system by the radiolysis of water molecules, leading to the X-ray-induced reduction of O2 to 2H2O at the trinuclear copper cluster (TNC) of the enzyme. In this work, 12 crystallographic structures of Thermus thermophilus HB27 multicopper oxidase (Tth-MCO) in holo, apo and Hg-bound forms and with different X-ray absorbed doses have been determined. In holo Tth-MCO structures with four Cu atoms, the proton-donor residue Glu451 involved in O2 reduction was found in a double conformation: Glu451a ( approximately 7 A from the TNC) and Glu451b ( approximately 4.5 A from the TNC). A positive peak of electron density above 3.5sigma in an Fo - Fc map for Glu451a O(2) indicates the presence of a carboxyl functional group at the side chain, while its significant absence in Glu451b strongly suggests a carboxylate functional group. In contrast, for apo Tth-MCO and in Hg-bound structures neither the positive peak nor double conformations were observed. Together, these observations provide the first structural evidence for a proton-relay mechanism in the MCO family and also support previous studies indicating that Asp106 does not provide protons for this mechanism. In addition, eight composite structures (Tth-MCO-C1-8) with different X-ray-absorbed doses allowed the observation of different O2-reduction states, and a total depletion of T2Cu at doses higher than 0.2 MGy showed the high susceptibility of this Cu atom to radiation damage, highlighting the importance of taking radiation effects into account in biochemical interpretations of an MCO structure.

X-ray-induced catalytic active-site reduction of a multicopper oxidase: structural insights into the proton-relay mechanism and O2-reduction states.,Serrano-Posada H, Centeno-Leija S, Rojas-Trejo SP, Rodriguez-Almazan C, Stojanoff V, Rudino-Pinera E Acta Crystallogr D Biol Crystallogr. 2015 Dec 1;71(Pt 12):2396-411. doi:, 10.1107/S1399004715018714. Epub 2015 Nov 26. PMID:26627648^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.