A unique respiratory adaptation in Drosophila independent of supercomplex formation.

Large assemblies of respiratory chain complexes, known as supercomplexes, are present in the mitochondrial membrane in mammals and yeast, as well as in some bacterial membranes. The formation of supercomplexes is thought to contribute to efficient electron transfer, stabilization of each enzyme complex, and inhibition of reactive oxygen species (ROS) generation. In this study, mitochondria from various organisms were solubilized with digitonin, and then the solubilized complexes were separated by blue native PAGE (BN-PAGE).

Heterogeneity between Two α Subunits of αβ Human Hemoglobin and O Binding Properties: Raman, H Nuclear Magnetic Resonance, and Terahertz Spectra.

Following a previous detailed investigation of the β subunit of αβ human adult hemoglobin (Hb A), this study focuses on the α subunit by using three natural valency hybrid α(Fe-deoxy/O)β(Fe) hemoglobin M (Hb M) in which O cannot bind to the β subunit: Hb M Hyde Park (β92His → Tyr), Hb M Saskatoon (β63His → Tyr), and Hb M Milwaukee (β67Val → Glu). In contrast with the β subunit that exhibited a clear correlation between O affinity and Fe-His stretching frequencies, the Fe-His stretching mode of the α subunit gave two Raman bands only in the T quaternary structure. This means the presence of two tertiary structures in α subunits of the αβ tetramer with T structure, and the two structures seemed to be nondynamical as judged from terahertz absorption spectra in the 5-30 cm region of Hb M Milwaukee, α(Fe-deoxy)β(Fe).

Activation Mechanism of the Streptomyces Tyrosinase Assisted by the Caddie Protein.

Tyrosinase (EC 1.14.18.

Rational Design of Domain-Swapping-Based c-Type Cytochrome Heterodimers by Using Chimeric Proteins.

The design of protein oligomers with multiple active sites has been gaining interest, owing to their potential use for biomaterials, which has encouraged researchers to develop a new design method. Three-dimensional domain swapping is the unique phenomenon in which protein molecules exchange the same structural region between each other. Herein, to construct oligomeric heme proteins with different active sites by utilizing domain swapping, two c-type cytochrome-based chimeric proteins have been constructed and the domains swapped.

A Ruthenium(III)-Oxyl Complex Bearing Strong Radical Character.

Proton-coupled electron-transfer oxidation of a Ru -OH complex, having an N-heterocyclic carbene ligand, gives a Ru -O species, which has an electronically equivalent structure of the Ru =O species, in an acidic aqueous solution. The Ru -O complex was characterized by spectroscopic methods and DFT calculations. The oxidation state of the Ru center was shown to be close to +3; the Ru-O bond showed a lower-energy Raman scattering at 732 cm and the Ru-O bond length was estimated to be 1.

Heme Orientation of Cavity Mutant Hemoglobins (His F8 → Gly) in Either α or β Subunits: Circular Dichroism, (1) H NMR, and Resonance Raman Studies.

Native human adult hemoglobin (Hb A) has mostly normal orientation of heme, whereas recombinant Hb A (rHb A) expressed in E. coli contains both normal and reversed orientations of heme. Hb A with the normal heme exhibits positive circular dichroism (CD) bands at both the Soret and 260-nm regions, while rHb A with the reversed heme shows a negative Soret and decreased 260-nm CD bands.

The functional role of the structure of the dioxo-isobacteriochlorin in the catalytic site of cytochrome cd for the reduction of nitrite.

Cytochrome cd is a key enzyme in bacterial denitrification and catalyzes one-electron reduction of nitrite (NO) to nitric oxide (NO) at the heme d center under anaerobic conditions. The heme d has a unique dioxo-isobacteriochlorin structure and is present only in cytochrome cd. To reveal the functional role of the unique heme d in the catalytic nitrite reduction, we studied effect of the porphyrin macrocycle on each reaction step of the catalytic cycle of cytochrome cd using synthetic model complexes.

A Model for the Active-Site Formation Process in DMSO Reductase Family Molybdenum Enzymes Involving Oxido-Alcoholato and Oxido-Thiolato Molybdenum(VI) Core Structures.

New bis(ene-1,2-dithiolato)-oxido-alcoholato molybdenum(VI) and -oxido-thiolato molybdenum(VI) anionic complexes, denoted as [Mo(VI)O(ER)L2](-) (E = O, S; L = dimethoxycarboxylate-1,2-ethylenedithiolate), were obtained from the reaction of the corresponding dioxido-molybdenum(VI) precursor complex with either an alcohol or a thiol in the presence of an organic acid (e.g., 10-camphorsulfonic acid) at low temperature.

Effects of Heme Electronic Structure and Distal Polar Interaction on Functional and Vibrational Properties of Myoglobin.

We analyzed the oxygen (O2) and carbon monoxide (CO) binding properties, autoxidation reaction rate, and FeO2 and FeCO vibrational frequencies of the H64Q mutant of sperm whale myoglobin (Mb) reconstituted with chemically modified heme cofactors possessing a variety of heme Fe electron densities (ρ(Fe)), and the results were compared with those for the previously studied native [Shibata, T. et al. J.

An Origin of Cooperative Oxygen Binding of Human Adult Hemoglobin: Different Roles of the α and β Subunits in the α2β2 Tetramer.

Human hemoglobin (Hb), which is an α2β2 tetramer and binds four O2 molecules, changes its O2-affinity from low to high as an increase of bound O2, that is characterized by 'cooperativity'. This property is indispensable for its function of O2 transfer from a lung to tissues and is accounted for in terms of T/R quaternary structure change, assuming the presence of a strain on the Fe-histidine (His) bond in the T state caused by the formation of hydrogen bonds at the subunit interfaces. However, the difference between the α and β subunits has been neglected.

Effects of proton motive force on the structure and dynamics of bovine cytochrome C oxidase in phospholipid vesicles.

A conventional method for reconstituting cytochrome c oxidase (CcO) into phospholipid vesicles (COV) has been modified to permit resonance Raman (RR) analysis in the presence and absence of proton motive force (ΔμH(+)). The COV has an average diameter of 20 nm and contains one CcO molecule within a unified orientation with CuA located outside the COV. The process of generation of ΔμH(+) across the membrane was monitored spectrophotometrically with rhodamine123 dye.

Infrared and Raman spectroscopic investigation of the reaction mechanism of cytochrome c oxidase.

Recent progress in studies on the proton-pumping and O₂reduction mechanisms of cytochrome c oxidase (CcO) elucidated by infrared (IR) and resonance Raman (rR) spectroscopy, is reviewed. CcO is the terminal enzyme of the respiratory chain and its O₂reduction reaction is coupled with H⁺ pumping activity across the inner mitochondrial membrane. The former is catalyzed by heme a3 and its mechanism has been determined using a rR technique, while the latter used the protein moiety and has been investigated with an IR technique.