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BioMed Research International 2021Cytochrome (CYP) enzymes catalyze the metabolic reactions of endogenous and exogenous compounds. The superfamily of enzymes is found across many organisms, regardless of...
Cytochrome (CYP) enzymes catalyze the metabolic reactions of endogenous and exogenous compounds. The superfamily of enzymes is found across many organisms, regardless of type, except for plants. Information was gathered about CYP2D enzymes through protein sequences of humans and other organisms. The secondary structure was predicted using the SOPMA. The structural and functional study of human CYP2D was conducted using ProtParam, SOPMA, Predotar 1.03, SignalP, TMHMM 2.0, and ExPASy. Most animals shared five central motifs according to motif analysis results. The tertiary structure of human CYP2D, as well as other animal species, was predicted by Phyre2. Human CYP2D proteins are heavily conserved across organisms, according to the findings. This indicates that they are descended from a single ancestor. They calculate the ratio of alpha-helices to extended strands to beta sheets to random coils. Most of the enzymes are alpha-helix, but small amounts of the random coil were also found. The data were obtained to provide us with a better understanding of mammalian proteins' functions and evolutionary relationships.
Topics: Amino Acid Sequence; Animals; Computational Biology; Computer Simulation; Cytochrome P-450 Enzyme System; Cytochromes; Humans; Ligands; Mice; Models, Molecular; Phylogeny; Protein Conformation, alpha-Helical; Protein Interaction Domains and Motifs; Protein Structure, Secondary; Proteins; Sequence Alignment; Software
PubMed: 34046497
DOI: 10.1155/2021/5574789 -
Microbiology Spectrum Dec 2022The sulfate-reducing microbe Desulfovibrio ferrophilus is of interest due to its relatively rare ability to also grow with Fe(III) oxide as an electron acceptor and its...
The sulfate-reducing microbe Desulfovibrio ferrophilus is of interest due to its relatively rare ability to also grow with Fe(III) oxide as an electron acceptor and its rapid corrosion of metallic iron. Previous studies have suggested multiple agents for extracellular electron exchange including a soluble electron shuttle, electrically conductive pili, and outer surface multiheme -type cytochromes. However, the previous lack of a strategy for genetic manipulation of limited mechanistic investigations. We developed an electroporation-mediated transformation method that enabled replacement of genes of interest with an antibiotic resistance gene via double-crossover homologous recombination. Genes were identified that are essential for flagellum-based motility and the expression of the two types of pili. Disrupting flagellum-based motility or expression of either of the two pili did not inhibit Fe(III) oxide reduction, nor did deleting genes for multiheme -type cytochromes predicted to be associated with the outer membrane. Although redundancies in cytochrome or pilus function might explain some of these phenotypes, overall, the results are consistent with primarily reducing Fe(III) oxide via an electron shuttle. The finding that is genetically tractable not only will aid in elucidating further details of its mechanisms for Fe(III) oxide reduction but also provides a new experimental approach for developing a better understanding of some of its other unique features, such as the ability to corrode metallic iron at high rates and accept electrons from negatively poised electrodes. is an important pure culture model for Fe(III) oxide reduction and the corrosion of iron-containing metals in anaerobic marine environments. This study demonstrates that is genetically tractable, an important advance for elucidating the mechanisms by which it interacts with extracellular electron acceptors and donors. The results demonstrate that there is not one specific outer surface multiheme -type cytochrome that is essential for Fe(III) oxide reduction. This finding, coupled with the lack of apparent porin-cytochrome conduits encoded in the genome and the finding that deleting genes for pilus and flagellum expression did not inhibit Fe(III) oxide reduction, suggests that has adopted strategies for extracellular electron exchange that are different from those of intensively studied electroactive microbes like and species. Thus, the ability to genetically manipulate is likely to lead to new mechanistic concepts in electromicrobiology.
Topics: Oxides; Oxidation-Reduction; Electron Transport; Ferric Compounds; Cytochromes; Iron
PubMed: 36445123
DOI: 10.1128/spectrum.03922-22 -
FEBS Letters Apr 2002Cytochrome cM is a new c-class photosynthetic haem protein whose physiological role is still unknown. It has been proposed previously that cytochrome cM can replace... (Comparative Study)
Comparative Study
Cytochrome cM is a new c-class photosynthetic haem protein whose physiological role is still unknown. It has been proposed previously that cytochrome cM can replace cytochrome c6 and plastocyanin in transferring electrons between the two membrane complexes cytochrome b6-f and photosystem I in organisms growing under stress conditions. The experimental evidence herein provided allows us to discard such a hypothesis. We report a procedure to overexpress cytochrome cM from the cyanobacterium Synechocystis sp. PCC 6803 in Escherichia coli cells in mg quantities. This has allowed us to perform a comparative laser flash-induced kinetic analysis of photosystem I reduction by the three metalloproteins from Synechocystis. The bimolecular rate constant for the overall reaction is up to 100 times lower with cytochrome cM than with cytochrome c6 or plastocyanin. In addition, the redox potential value and surface electrostatic potential distribution of cytochrome cM are quite different from those of cytochrome c6 and plastocyanin. These findings strongly indicate that cytochrome cM cannot be recognised by and interact with the same redox partners as the other two metalloproteins.
Topics: Amino Acid Sequence; Cyanobacteria; Cytochromes; Cytochromes f; Electron Transport; Models, Molecular; Molecular Sequence Data; Oxidation-Reduction; Photosynthetic Reaction Center Complex Proteins; Photosystem I Protein Complex; Plastocyanin; Protein Conformation; Sequence Alignment; Static Electricity
PubMed: 12062408
DOI: 10.1016/s0014-5793(02)02576-0 -
British Journal of Pharmacology Mar 2008The cytochromes P450 (CYPs) comprise a vast superfamily of enzymes found in virtually all life forms. In mammals, xenobiotic metabolizing CYPs provide crucial protection... (Review)
Review
The cytochromes P450 (CYPs) comprise a vast superfamily of enzymes found in virtually all life forms. In mammals, xenobiotic metabolizing CYPs provide crucial protection from the effects of exposure to a wide variety of chemicals, including environmental toxins and therapeutic drugs. Ideally, the information on the possible metabolism by CYPs required during drug development would be obtained from crystal structures of all the CYPs of interest. For some years only crystal structures of distantly related bacterial CYPs were available and homology modelling techniques were used to bridge the gap and produce structural models of human CYPs, and thereby obtain useful functional information. A significant step forward in the reliability of these models came seven years ago with the first crystal structure of a mammalian CYP, rabbit CYP2C5, followed by the structures of six human enzymes, CYP1A2, CYP2A6, CYP2C8, CYP2C9, CYP2D6 and CYP3A4, and a second rabbit enzyme, CYP2B4. In this review we describe as a case study the evolution of a CYP2D6 model, leading to the validation of the model as an in silico tool for predicting binding and metabolism. This work has led directly to the successful design of CYP2D6 mutants with novel activity-including creating a testosterone hydroxylase, converting quinidine from inhibitor to substrate, creating a diclofenac hydroxylase and creating a dextromethorphan O-demethylase. Our modelling-derived hypothesis-driven integrated interdisciplinary studies have given key insight into the molecular determinants of CYP2D6 and other important drug metabolizing enzymes.
Topics: Animals; Cytochrome P-450 CYP2D6; Cytochrome P-450 Enzyme System; Drug Interactions; Humans; Models, Molecular; Pharmaceutical Preparations; Substrate Specificity
PubMed: 18026129
DOI: 10.1038/sj.bjp.0707570 -
The Journal of Biological Chemistry Aug 1983The aerobic respiratory chain of Escherichia coli is branched. In aerobically grown cells harvested in midexponential phase, a respiratory chain containing only b-type...
The aerobic respiratory chain of Escherichia coli is branched. In aerobically grown cells harvested in midexponential phase, a respiratory chain containing only b-type cytochromes is predominant. This chain contains a terminal oxidase which is a b-type cytochrome, referred to as cytochrome o. However, when the bacteria are grown under conditions of oxygen limitation, additional components of the respiratory chain are induced, as evidenced by the appearance of new spectroscopic species. These include a new b-type cytochrome, cytochrome b558, as well as cytochrome a1 and cytochrome d. In this paper, a purification protocol and the initial characterization of the terminal oxidase complex containing cytochrome d are reported. Solubilization of the membrane is effected by Zwittergent 3-12, and purification is accomplished by chromatography with DEAE-Sepharose CL-6B and hydroxyapatite. The complex contains cytochrome b558, a1, and d. Analysis by sodium dodecyl sulfate-polyacrylamide gels indicates that the complex contains only two types of polypeptides with the molecular weights estimated to be 57,000 and 43,000. The purified complex has oxidase activity in the presence of detergents, utilizing substrates including ubinquinol-1, N,N,N',N'-tetramethyl-p-phenylenediamine, and 2,3,5,6-tetramethyl-p-phenylenediamine. The cytochrome d complex contains protoheme IX and iron, but does not contain nonheme iron or copper. Approximately half of the cytochromes which are thought to participate in E. coli aerobic respiration are accounted for by this single complex. These results suggest that the E. coli aerobic respiratory chain is organized around a relatively small number of cytochrome-containing complexes.
Topics: Cytochrome b Group; Cytochrome d Group; Cytochromes; Cytochromes a1; Electron Spin Resonance Spectroscopy; Escherichia coli; Escherichia coli Proteins; Macromolecular Substances; Molecular Weight; NADPH Oxidases; Spectrophotometry
PubMed: 6307994
DOI: No ID Found -
The Journal of Biological Chemistry Mar 1984Cytochrome b558-d complex, a terminal oxidase in the respiratory chain of Escherichia coli K12 grown under the condition of limited oxygen, was purified to near...
Terminal oxidases of Escherichia coli aerobic respiratory chain. II. Purification and properties of cytochrome b558-d complex from cells grown with limited oxygen and evidence of branched electron-carrying systems.
Cytochrome b558-d complex, a terminal oxidase in the respiratory chain of Escherichia coli K12 grown under the condition of limited oxygen, was purified to near homogeneity. The purified oxidase complex is composed of equimolar amounts of two polypeptides, with Mr = 26,000 and 51,000, determined with gel electrophoresis in the presence of sodium dodecyl sulfate. It contains 12.3 nmol of protoheme, 9.54 nmol of cytochrome d, and 26.6 nmol of iron/mg of protein. The enzyme is a "cytochrome bd-type oxidase," which shows absorption peaks at 558 and 624 nm in the difference spectrum at 77 K. This oxidase combines with CO, and the Soret region of the CO difference spectrum at room temperature has a peak at 420 nm and troughs at 430 and 442 nm. The oxidation-reduction potentials of cytochrome b558 and cytochrome d at pH 7.4 were estimated to be +10 mV and +240 mV, respectively. The cytochrome b558-d complex catalyzes the oxidation of ubiquinol-1, menadiol, and ascorbate in the presence of N,N,N',N'-tetramethyl-p-phenylenediamine dihydrochloride. This oxidase activity was inhibited by the respiratory inhibitors piericidin A, KCN, and NaN3, but the cytochrome b558-d complex was less sensitive to the inhibitors than the cytochrome b562-o complex. The Km values for oxygen of purified cytochrome b558-d complex and cytochrome b562-o complex were determined to be 0.38 and 2.9 microM, respectively. Formation of a membrane potential by the reconstituted cytochrome b558-d complex in liposomes was observed with the fluorescent dye 3,3'-dipropylthiocarbocyanine iodide on addition of ubiquinol-1. This is the first indication that there is a coupling site in the terminal oxidase, which contains cytochrome d. Steady state kinetics of cytochromes in the membrane showed that these oxidase complexes branch at the site of ubiquinone-8 in the respiratory chain. From these and previous results, branched electron-carrying systems of the E. coli respiratory chain are proposed.
Topics: Aerobiosis; Cell Membrane; Cytochrome b Group; Cytochrome d Group; Cytochromes; Electron Transport; Escherichia coli; Kinetics; Multienzyme Complexes; NADPH Oxidases; Oxidation-Reduction; Oxygen Consumption; Potentiometry; Quinone Reductases; Spectrophotometry
PubMed: 6321507
DOI: No ID Found -
Membrane environment drives cytochrome P450's spin transition and its interaction with cytochrome b.Chemical Communications (Cambridge,... Nov 2017Heme's spin-multiplicity is key in determining the enzymatic function of cytochrome P450 (cytP450). The origin of the low-spin state in ferric P450 is still under...
Heme's spin-multiplicity is key in determining the enzymatic function of cytochrome P450 (cytP450). The origin of the low-spin state in ferric P450 is still under debate. Here, we report the first experimental demonstration of P450's membrane interaction altering its spin equilibrium which is accompanied by a stronger affinity for cytochrome b. These results highlight the importance of lipid membrane for the function of P450.
Topics: Cytochrome P-450 Enzyme System; Cytochromes b5; Models, Molecular
PubMed: 29143058
DOI: 10.1039/c7cc07520k -
Biochimica Et Biophysica Acta Sep 2016Cytochrome cd1 nitrite reductases (cd1NiRs) catalyze the reduction of nitrite to nitric oxide in denitrifying bacteria, such as Marinobacter hydrocarbonoclasticus.... (Comparative Study)
Comparative Study
Cytochrome cd1 nitrite reductases (cd1NiRs) catalyze the reduction of nitrite to nitric oxide in denitrifying bacteria, such as Marinobacter hydrocarbonoclasticus. Previous work demonstrated that the enzymatic activity depends on a structural pre-activation triggered by the entry of electrons through the electron transfer (ET) domain, which houses a heme c center. The catalytic activity of M. hydrocarbonoclasticus cd1NiR (Mhcd1NiR) was tested by mediated electrochemistry, using small ET proteins and chemical redox mediators. The rate of enzymatic reaction depends on the nature of the redox partner, with cytochrome (cyt) c552 providing the highest value. In situations where cyt c552 is replaced by either a biological (cyt c from horse heart) or a chemical mediator the catalytic response was only observed at very low scan rates, suggesting that the intermolecular ET rate is much slower. Molecular docking simulations with the 3D model structure of Mhcd1NiR and cyt c552 or cyt c showed that hydrophobic interactions favor the formation of complexes where the heme c domain of the enzyme is the principal docking site. However, only in the case of cyt c552 the preferential areas of contact and Fe-Fe distances between heme c groups of the redox partners allow establishing competent ET pathways. The coupling of the enzyme with chemical redox mediators was also found not to be energetically favorable. These results indicate that although low activity functional complexes can be formed between Mhcd1NiR and different types of redox mediators, efficient ET is only observed with the putative physiological electron donor cyt c552.
Topics: Cytochrome c Group; Cytochromes; Electron Transport; Molecular Docking Simulation; Nitrite Reductases; Oxidation-Reduction
PubMed: 27133504
DOI: 10.1016/j.bbabio.2016.04.279 -
Structure (London, England : 1993) Feb 1994Cytochrome f is the high potential electron acceptor of the chloroplast cytochrome b6f complex, and is the electron donor to plastocyanin. The 285-residue cytochrome f... (Comparative Study)
Comparative Study
BACKGROUND
Cytochrome f is the high potential electron acceptor of the chloroplast cytochrome b6f complex, and is the electron donor to plastocyanin. The 285-residue cytochrome f subunit is anchored in the thylakoid membrane of the chloroplast by a single membrane-spanning segment near the carboxyl terminus. A soluble redox-active 252-residue lumen-side polypeptide with native spectroscopic and redox properties, missing the membrane anchor and carboxyl terminus, was purified from turnip chloroplasts for structural studies.
RESULTS
The crystal structure of cytochrome f, determined to 2.3 A resolution, has several unexpected features. The 252-residue polypeptide is organized into one large and one small domain. The larger heme-binding domain is strikingly different from known structures of other c-type cytochromes and has the same fold as the type III domain of the animal protein, fibronectin. Cytochrome f binds heme with an unprecedented axial heme iron ligand: the amino terminus of the polypeptide.
CONCLUSION
The first atomic structure of a subunit of either the cytochrome b6f complex or of the related cytochrome bc1 complex has been obtained. The structure of cytochrome f allows prediction of the approximate docking site of plastocyanin and should allow systematic studies of the mechanism of intra- and inter-protein electron transfer between the cytochrome heme and plastocyanin copper, which are approximately isopotential. The unprecedented axial heme iron ligand also provides information on the sequence of events (i.e. cleavage of signal peptide and ligation of heme) associated with translocation of the cytochrome across the membrane and its subsequent folding.
Topics: Amino Acid Sequence; Binding Sites; Chloroplasts; Crystallography, X-Ray; Cytochromes; Cytochromes f; Heme; Models, Molecular; Molecular Sequence Data; Protein Folding; Protein Structure, Secondary; Vegetables
PubMed: 8081747
DOI: 10.1016/s0969-2126(00)00012-5 -
European Journal of Biochemistry Mar 2001Using 1617 meaningful NOEs with 188 pseudocontact shifts, a family of 35 conformers of oxidized bovine microsomal cytochrome b5 mutant (E44/48/56A/D60A) has been...
Using 1617 meaningful NOEs with 188 pseudocontact shifts, a family of 35 conformers of oxidized bovine microsomal cytochrome b5 mutant (E44/48/56A/D60A) has been obtained and is characterized by good resolution (rmsd to the mean structure are 0.047 +/- 0.007 nm and 0.095 +/- 0.008 nm for backbone and heavy atoms, respectively). The solution structure of the mutant, when compared with the X-ray structure of wild-type cytochrome b(5), has no significant changes in the whole folding and secondary structure. The binding between cytochrome b(5) and cytochrome c shows that the association constant of the mutant-cytochrome c complex is much lower than the one for wild-type complex (2.2 x 10(4) M(-1) vs. 5.1 x 10(3) M(-1)). The result suggests the four acidic residues have substantial effects on the formation of the complex between cytochrome b(5) and cytochrome c, and therefore it is concluded reasonably that the electrostatic interaction plays an important role in maintaining the stability and specificity of the complex formed. The competition between the ferricytochrome b(5) mutant and [Cr(oxalate)(3)](3-) for ferricytochrome c shows that site III of cytochrome c, which is a strong binding site to wild-type cytochrome b(5), still binds to the mutant with relatively weaker strength. Our results indicate that certain bonding geometries do occur in the interaction between the present mutant and cytochrome c and these geometries, which should be quite different from the ones of the Salemme and Northrup models.
Topics: Amino Acid Sequence; Cytochrome c Group; Cytochromes b5; Magnetics; Models, Molecular; Molecular Sequence Data; Mutation; Nuclear Magnetic Resonance, Biomolecular; Protein Conformation
PubMed: 11248680
DOI: No ID Found