Abstract. Cytochrome c peroxidase (CCP) is a nuclearly encoded hemoprotein located in the intermembrane space (IMS) of Saccharomyces cerevisiae mitochondria. Wild-type preCCP synthesized in rabbit reticulocyte lysates, however, was inefficiently translocated into isolated mitochondria and was inherently resistant to externally added proteases. To test whether premature heme addition to the apoprecursor was responsible for the protease resistance and the inability to import preCCP, site-directed mutagenesis was used to replace the axial heme ligand (Hiss75) involved in forming a pseudo-covalent link between the heme iron and CCP. Mutant proteins containing Leu, Arg, Met, or Pro at residue 175 of mature CCP were sensitive to proteolysis and were imported into isolated mitochondria as judged by proteolytic processing of the precursor. The inhibition of wild-type CCP translocation across the outer membrane may result from the inability of the heme-containing protein to unfold during the translocation process.Although the protease responsible for cleaving preCCP to its mature form is believed to be located in the IMS, most of the processed CCP was located in the supernatant rather than the mitochondrial pellet. Since the outer membranes were shown to be intact, the anomalous localization indicated that preCCP may not have been completely translocated into the IMS before proteolytic processing or that conformationally labile proteins may not be retained by the outer membrane. Proteolytic maturation of preCCP also occurred in the presence of valinomycin, suggesting that the precursor may be completely or partially translocated across the outer mitochondrial membrane independent of a potential across the inner mitochondrial membrane. MOST mitochondrial proteins are encoded in the nucleus, synthesized as larger precursors in the cytoplasm (37), and then specifically targeted to one of four mitochondrial compartments: the outer membrane, the intermembrane space (IMS),~ the inner membrane, or the Portions of this work have appeared in abstract form (Brandriss, M. C., K.

The reactions of C 3 v Ir3(μ-CO)3(η5-C9H7)3 (1) with various metal electrophiles yield the cationic tetranuclear clusters [Ir3{M(PPh3)}(CO)3(η5-C9H7)3][PF6] (2, M = Cu; 3, M = Ag; 5, M = Au), [Ir3Tl(μ-CO)3(η5-C9H7)3][PF6] (4), and [Ir3(HgR)(CO)3(η5-C9H7)3][PF6] (6, R = Ph; 7, R = W(CO)3(η5-C5H5)). Compounds 5−7 are best prepared via compound 4. The structures of compounds 4 and 5 have been determined by X-ray diffraction. The C 3 v symmetry of the parent cluster 1 is maintained in the structure of 4. The molecule consists of a triangle of iridium atoms, each edge of which has a bridging carbonyl oriented out of the plane in the same direction and each vertex of which has an η5-indenyl ligand oriented toward the opposite side of the plane. The thallium atom adopts a face-capping mode of coordination on the same side of the triiridium plane as the three indenyl ligands and is encapsulated by the phenylene portions of the indenyl groups. The molecular structure of 5 consists of a AuIr3 butterfly framework with a hinge angle of 153.63(3)° and the gold atom in a wingtip position. Each CO ligand is bonded in a terminal mode to one iridium center, with one CO ligand ‘down' relative to the Ir3 plane and two CO ligands ‘up', flanking the Ir−Ir edge bridged by the Au(PPh3)+ fragment. The indenyl ligands take up positions opposite those of the CO ligands at each iridium center. The infrared and 1H NMR spectra of compounds 2, 3, 5, 6, and 7 are all very similar and are fully consistent with the solid-state structure of 5.

The slow addition of Re(2)(&mgr;-H)(2)(CO)(8) to a solution of Ir(CO)(eta(2)-C(8)H(14))(eta(5)-C(9)H(7)) in hexane at reflux provides IrRe(2)(&mgr;-H)(2)(CO)(9)(eta(5)-C(9)H(7)) (1) in 80% yield. The molecular structure of 1 shows an IrRe(2) triangle incorporating one Ir(CO)(eta(5)-C(9)H(7)) and two Re(CO)(4) fragments. The strongly different Ir-Re distances suggest that one hydride ligand bridges one Ir-Re edge and the other hydride bridges the Re-Re edge. Low-temperature (1)H and (13)C NMR spectra are consistent with this structure; at higher temperatures a dynamic process involving migration of one hydride ligand between the two Ir-Re edges is observed. Cluster 1 is readily deprotonated with KOH/EtOH, and the resulting anion has been isolated as the PPN salt, [PPN][IrRe(2)(&mgr;-H)(CO)(9)(eta(5)-C(9)H(7))] (2). Both the (1)H and low temperature (13)C NMR spectra of 2 are consistent with a structure in which the remaining hydride ligand bridges the Re-Re edge. Variable-temperature (13)C NMR spectra indicate that 2 undergoes CO scrambling localized on the Ir-Re edges. The reaction of 1 with PPh(3) leads to IrRe(2)(&mgr;-H)(2)(CO)(8)(PPh(3))(eta(5)-C(9)H(7)) (3), which contains the phosphine on a rhenium atom, as well as to cluster fragmentation.

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