The α proteobacter Rhodobacter sphaeroides accumulates two cytochrome c oxidases (CcO) in its cytoplasmic membrane during aerobic growth: a mitochondrial-like aa3-type CcO containing a di-copper CuA center and mono-copper CuB, plus a cbb3-type CcO that contains CuB but lacks CuA. Three copper chaperones are located in the periplasm of R. sphaeroides, PCuAC, PrrC (Sco) and Cox11. Cox11 is required to assemble CuB of the aa3-type but not the cbb3-type CcO. PrrC is homologous to mitochondrial Sco1; Sco proteins are implicated in CuA assembly in mitochondria and bacteria, and with CuB assembly of the cbb3-type CcO. PCuAC is present in many bacteria, but not mitochondria. PCuAC of Thermus thermophilus metallates a CuA center in vitro, but its in vivo function has not been explored. Here, the extent of copper center assembly in the aa3- and cbb3-type CcOs of R. sphaeroides has been examined in strains lacking PCuAC, PrrC, or both. The absence of either chaperone strongly lowers the accumulation of both CcOs in the cells grown in low concentrations of Cu2+. The absence of PrrC has a greater effect than the absence of PCuAC and PCuAC appears to function upstream of PrrC. Analysis of purified aa3-type CcO shows that PrrC has a greater effect on the assembly of its CuA than does PCuAC, and both chaperones have a lesser but significant effect on the assembly of its CuB even though Cox11 is present. Scenarios for the cellular roles of PCuAC and PrrC are considered. The results are most consistent with a role for PrrC in the capture and delivery of copper to CuA of the aa3-type CcO and to CuB of the cbb3-type CcO, while the predominant role of PCuAC may be to capture and deliver copper to PrrC and Cox11.

The Cu(I) chaperone Cox11 is required for the insertion of Cu B into cytochrome c oxidase (CcO) of mitochondria and many bacteria, including Rhodobacter sphaeroides. Exploration of the copper binding stoichiometry of R. sphaeroides Cox11 led to the finding that an apparent tetramer of both mitochondrial and bacterial Cox11 binds more copper than the sum of the dimers, providing another example of the flexibility of copper binding by Cu(I)-S clusters. Site-directed mutagenesis has been used to identify components of Cox11 that are not required for copper binding but are absolutely required for the assembly of Cu B , including conserved Cys-35 and Lys-123. In contrast to earlier proposals, Cys-35 is not required for dimerization of Cox11 or for copper binding. These findings, plus the location of Cys-35 at the C terminus of the predicted transmembrane helix and thereby close to the surface of the membrane, allows a proposal that Cys-35 is involved in the transfer of copper from the Cu(I) cluster of Cox11 to the Cu B ligands His-333/334 during the folding of CcO subunit I. Lys-123 is located near the Cu(I) cluster of Cox11, in an area otherwise devoid of charged residues. From the analysis of several Cox11 mutants, including K123E, L and R, we conclude that a previous proposal that Lys-123 provides charge balance for the stabilization of the Cu(I) cluster is unlikely to account for its absolute requirement for Cox11 function. Rather, consideration of the properties of Lys-123 and the apparent specificity of Cox11 suggests that Lys-123 plays a role in the interaction of Cox11 with its target.The aa 3 -type cytochrome c oxidase (CcO) is the terminal member of the respiratory electron transfer system in mitochondria and many aerobic bacteria (1,2). The CcO complex contains two separate copper centers, Cu A in subunit II and Cu B in subunit I. The Cu A center contains two copper ions, liganded by two bridging cysteines plus two histidines, one carboxyl side chain and one backbone carbonyl oxygen (3-5). Cu B is a component of the O 2 reduction site in subunit I, along with heme a 3 and a cross-linked His-Tyr cofactor (4-6). The single copper of Cu B is liganded by three histidines of subunit I, two of which are adjacent . The third ligand is the histidine of the His-Tyr cofactor. The structure of the Cu B center is highly conserved throughout the large heme-Cu oxidase superfamily (7). The structure of the Cu A center is also conserved, but this center is only present in certain groups of the heme-Cu oxidases. In bacteria, a CcO subcomplex containing only subunits I and II is readily produced (8,9). In this fully-folded and active I-II oxidase form, all of the redox centers, Cu A , heme a and the heme a 3 -Cu B center, are buried within the protein and inaccessible to solvent (10). The structure and stability of this complex predicts that all of the redox centers are assembled during the folding of these subunits in the membrane and prior to the association of subunit II with subunit I.The α-proteobacter Rhodobac...

In this study, hydrovanilloin synthesized by electrochemical dimerization of vanillin has been used as a renewable substitute for bisphenol A for the preparation of epoxy resins. The reaction of the disodium salt of hydrovanilloin:epichlorohydrin 1:2 mol ratio at 80°C for 30 min in water gave a hydrovanilloin – diglycidyl ether phenoxy resin. This hard thermoplastic resin showed Tg of 135°C and stable up to 255°C in air. On the other hand, the disodium salt of hydrovanilloin:epichlorohydrin 1:4 mol ratio at 80°C for 30 min in water gives a curable oligomer of hydrovanilloin – diglycidyl ether with 2.1 repeating units. This oligomer could be cured with aliphatic diamines: 1,2‐diaminoethane, 1,4‐diaminobutane, 1,6‐diaminohexane, and isophorone diamine to give hard epoxy resins with Tg values of 116, 118, 149 and 146°C, respectively. © 2018 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2019, 136, 47000.

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