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Biochimica Et Biophysica Acta Aug 2000Membrane-bound succinate dehydrogenases (succinate:quinone reductases, SQR) and fumarate reductases (quinol:fumarate reductases, QFR) couple the oxidation of succinate... (Review)
Review
Membrane-bound succinate dehydrogenases (succinate:quinone reductases, SQR) and fumarate reductases (quinol:fumarate reductases, QFR) couple the oxidation of succinate to fumarate to the reduction of quinone to quinol and also catalyse the reverse reaction. SQR (respiratory complex II) is involved in aerobic metabolism as part of the citric acid cycle and of the aerobic respiratory chain. QFR is involved in anaerobic respiration with fumarate as the terminal electron acceptor, and is part of an electron transport chain catalysing the oxidation of various donor substrates by fumarate. QFR and SQR complexes are collectively referred to as succinate:quinone oxidoreductases (EC 1.3.5.1), have very similar compositions and are predicted to share similar structures. The complexes consist of two hydrophilic and one or two hydrophobic, membrane-integrated subunits. The larger hydrophilic subunit A carries covalently bound flavin adenine dinucleotide and subunit B contains three iron-sulphur centres. QFR of Wolinella succinogenes and SQR of Bacillus subtilis contain only one hydrophobic subunit (C) with two haem b groups. In contrast, SQR and QFR of Escherichia coli contain two hydrophobic subunits (C and D) which bind either one (SQR) or no haem b group (QFR). The structure of W. succinogenes QFR has been determined at 2.2 A resolution by X-ray crystallography (C.R.D. Lancaster, A. Kröger, M. Auer, H. Michel, Nature 402 (1999) 377-385). Based on this structure of the three protein subunits and the arrangement of the six prosthetic groups, a pathway of electron transfer from the quinol-oxidising dihaem cytochrome b to the site of fumarate reduction and a mechanism of fumarate reduction was proposed. The W. succinogenes QFR structure is different from that of the haem-less QFR of E. coli, described at 3.3 A resolution (T.M. Iverson, C. Luna-Chavez, G. Cecchini, D.C. Rees, Science 284 (1999) 1961-1966), mainly with respect to the structure of the membrane-embedded subunits and the relative orientations of soluble and membrane-embedded subunits. Also, similarities and differences between QFR transmembrane helix IV and transmembrane helix F of bacteriorhodopsin and their implications are discussed.
Topics: Animals; Binding Sites; Crystallography, X-Ray; Electron Transport; Electron Transport Complex II; Escherichia coli; Flavoproteins; Humans; Iron-Sulfur Proteins; Membrane Potentials; Membrane Proteins; Models, Chemical; Models, Molecular; Molecular Structure; Multienzyme Complexes; Oxidoreductases; Succinate Dehydrogenase; Wolinella
PubMed: 11004459
DOI: 10.1016/s0005-2728(00)00180-8 -
Biomolecules Jun 2021The use of multienzyme complexes can facilitate biocatalytic cascade reactions by employing fusion enzymes or protein tags. In this study, we explored the use of...
The use of multienzyme complexes can facilitate biocatalytic cascade reactions by employing fusion enzymes or protein tags. In this study, we explored the use of recently developed peptide tags that promote complex formation of the targeted proteins: the dimerization-docking and anchoring domain (RIDD-RIAD) system. These peptides allow self-assembly based on specific protein-protein interactions between both peptides and allow tuning of the ratio of the targeted enzymes as the RIAD peptide binds to two RIDD peptides. Each of these tags were added to the C-terminus of a NADPH-dependent Baeyer-Villiger monooxygenase (phenylacetone monooxygenase, PAMO) and a NADPH-regenerating enzyme (phosphite dehydrogenase, PTDH). Several RIDD/RIAD-tagged PAMO and PTDH variants were successfully overproduced in and subsequently purified. Complementary tagged enzymes were mixed and analyzed for their oligomeric state, stability, and activity. Complexes were formed in the case of some specific combinations (PAMO-PTDH and PAMO-PTDH). These enzyme complexes displayed similar catalytic activity when compared with the PTDH-PAMO fusion enzyme. The thermostability of PAMO in these complexes was retained while PTDH displayed somewhat lower thermostability. Evaluation of the biocatalytic performance by conducting conversions revealed that with a self-assembled PAMO-PTDH complex less PTDH was required for the same performance when compared with the PTDH-PAMO fusion enzyme.
Topics: Mixed Function Oxygenases; Multienzyme Complexes; NADH, NADPH Oxidoreductases; Recombinant Proteins
PubMed: 34204515
DOI: 10.3390/biom11060905 -
Biochimica Et Biophysica Acta Jan 2002
Review
Topics: Bacteria; Benzoquinones; Electron Transport Complex II; Fumarates; Hydroquinones; Intracellular Membranes; Models, Chemical; Models, Molecular; Multienzyme Complexes; Oxidoreductases; Succinate Dehydrogenase
PubMed: 11803013
DOI: 10.1016/s0005-2728(01)00240-7 -
Cellular and Molecular Life Sciences :... Mar 1998The barrel-shaped 20S proteasome is one of the two components of a larger 26S particle, the multicatalytic 2000-kDa protease complex. The proteolytic sites are located... (Review)
Review
The barrel-shaped 20S proteasome is one of the two components of a larger 26S particle, the multicatalytic 2000-kDa protease complex. The proteolytic sites are located in the inner chamber of the 20S particle and are only accessible via narrow entrances. This paper reviews the current knowledge concerning proteasome formation, proteolytic activities, structural aspects and assembly. Eukaryotic proteasomes are made up by four rings each of which contains seven different subunits occurring at fixed positions. While the outer rings contain alpha-type subunits, the inner ones comprise beta-type subunits. The current assembly model for eukaryotic 20S proteasomes is based upon the detection of 13S and 16S intermediates, respectively, in addition to previous findings with archaebacterial and eubacterial proteasome assembly. The available data suggest a cooperative assembly of the alpha-type and beta-type subunits into half proteasome-like complexes followed by dimerization into proteasomes. During or after dimerization of half proteasomes, the beta-type subunits are processed. The prosequence of the beta-type subunits is essential for the assembly proves and prevents protease activity of immature proteasomes.
Topics: Amino Acid Sequence; Bacterial Proteins; Cysteine Endopeptidases; Humans; Hydrolysis; Molecular Sequence Data; Multienzyme Complexes; Proteasome Endopeptidase Complex; Sequence Homology, Amino Acid
PubMed: 9575337
DOI: 10.1007/s000180050147 -
Biochemistry May 2014Two hallmarks of assembly line polyketide synthases have motivated an interest in these unusual multienzyme systems, their stereospecificity and their capacity for... (Review)
Review
Two hallmarks of assembly line polyketide synthases have motivated an interest in these unusual multienzyme systems, their stereospecificity and their capacity for directional biosynthesis. In this review, we summarize the state of knowledge regarding the mechanistic origins of these two remarkable features, using the 6-deoxyerythronolide B synthase as a prototype. Of the 10 stereocenters in 6-deoxyerythronolide B, the stereochemistry of nine carbon atoms is directly set by ketoreductase domains, which catalyze epimerization and/or diastereospecific reduction reactions. The 10th stereocenter is established by the sequential action of three enzymatic domains. Thus, the problem has been reduced to a challenge in mainstream enzymology, where fundamental gaps remain in our understanding of the structural basis for this exquisite stereochemical control by relatively well-defined active sites. In contrast, testable mechanistic hypotheses for the phenomenon of vectorial biosynthesis are only just beginning to emerge. Starting from an elegant theoretical framework for understanding coupled vectorial processes in biology [Jencks, W. P. (1980) Adv. Enzymol. Relat. Areas Mol. Biol. 51, 75-106], we present a simple model that can explain assembly line polyketide biosynthesis as a coupled vectorial process. Our model, which highlights the important role of domain-domain interactions, not only is consistent with recent observations but also is amenable to further experimental verification and refinement. Ultimately, a definitive view of the coordinated motions within and between polyketide synthase modules will require a combination of structural, kinetic, spectroscopic, and computational tools and could be one of the most exciting frontiers in 21st Century enzymology.
Topics: Models, Chemical; Models, Molecular; Multienzyme Complexes; Polyketide Synthases; Protein Structure, Tertiary; Stereoisomerism
PubMed: 24779441
DOI: 10.1021/bi500290t -
Biochimica Et Biophysica Acta Jan 2002Succinate-ubiquinone oxidoreductase (SQR) as part of the trichloroacetic acid cycle and menaquinol-fumarate oxidoreductase (QFR) used for anaerobic respiration by... (Comparative Study)
Comparative Study Review
Succinate-ubiquinone oxidoreductase (SQR) as part of the trichloroacetic acid cycle and menaquinol-fumarate oxidoreductase (QFR) used for anaerobic respiration by Escherichia coli are structurally and functionally related membrane-bound enzyme complexes. Each enzyme complex is composed of four distinct subunits. The recent solution of the X-ray structure of QFR has provided new insights into the function of these enzymes. Both enzyme complexes contain a catalytic domain composed of a subunit with a covalently bound flavin cofactor, the dicarboxylate binding site, and an iron-sulfur subunit which contains three distinct iron-sulfur clusters. The catalytic domain is bound to the cytoplasmic membrane by two hydrophobic membrane anchor subunits that also form the site(s) for interaction with quinones. The membrane domain of E. coli SQR is also the site where the heme b556 is located. The structure and function of SQR and QFR are briefly summarized in this communication and the similarities and differences in the membrane domain of the two enzymes are discussed.
Topics: Binding Sites; Cell Membrane; Electron Transport Complex II; Escherichia coli; Gene Expression Regulation, Bacterial; Gene Expression Regulation, Enzymologic; Hydrogen-Ion Concentration; Intracellular Membranes; Iron-Sulfur Proteins; Kinetics; Models, Molecular; Multienzyme Complexes; Multigene Family; Oxidoreductases; Structure-Activity Relationship; Succinate Dehydrogenase
PubMed: 11803023
DOI: 10.1016/s0005-2728(01)00238-9 -
Cell Sep 2017The respiratory megacomplex represents the highest-order assembly of respiratory chain complexes, and it allows mitochondria to respond to energy-requiring conditions....
The respiratory megacomplex represents the highest-order assembly of respiratory chain complexes, and it allows mitochondria to respond to energy-requiring conditions. To understand its architecture, we examined the human respiratory chain megacomplex-IIIIIV (MCIIIIIV) with 140 subunits and a subset of associated cofactors using cryo-electron microscopy. The MCIIIIIV forms a circular structure with the dimeric CIII located in the center, where it is surrounded by two copies each of CI and CIV. Two cytochrome c (Cyt.c) molecules are positioned to accept electrons on the surface of the c state CIII dimer. Analyses indicate that CII could insert into the gaps between CI and CIV to form a closed ring, which we termed the electron transport chain supercomplex. The structure not only reveals the precise assignment of individual subunits of human CI and CIII, but also enables future in-depth analysis of the electron transport chain as a whole.
Topics: Cryoelectron Microscopy; Electron Transport Chain Complex Proteins; Electron Transport Complex I; Electron Transport Complex II; Humans; Mitochondria; Models, Molecular; Multienzyme Complexes
PubMed: 28844695
DOI: 10.1016/j.cell.2017.07.050 -
Nature Structural & Molecular Biology Dec 2015At the eukaryotic DNA replication fork, it is widely believed that the Cdc45-Mcm2-7-GINS (CMG) helicase is positioned in front to unwind DNA and that DNA polymerases...
At the eukaryotic DNA replication fork, it is widely believed that the Cdc45-Mcm2-7-GINS (CMG) helicase is positioned in front to unwind DNA and that DNA polymerases trail behind the helicase. Here we used single-particle EM to directly image a Saccharomyces cerevisiae replisome. Contrary to expectations, the leading strand Pol ɛ is positioned ahead of CMG helicase, whereas Ctf4 and the lagging-strand polymerase (Pol) α-primase are behind the helicase. This unexpected architecture indicates that the leading-strand DNA travels a long distance before reaching Pol ɛ, first threading through the Mcm2-7 ring and then making a U-turn at the bottom and reaching Pol ɛ at the top of CMG. Our work reveals an unexpected configuration of the eukaryotic replisome, suggests possible reasons for this architecture and provides a basis for further structural and biochemical replisome studies.
Topics: DNA Replication; DNA, Fungal; Image Processing, Computer-Assisted; Microscopy, Electron; Multienzyme Complexes; Saccharomyces cerevisiae
PubMed: 26524492
DOI: 10.1038/nsmb.3113 -
Microbiology and Molecular Biology... Dec 2009The Mcm2-7 complex serves as the eukaryotic replicative helicase, the molecular motor that both unwinds duplex DNA and powers fork progression during DNA replication.... (Review)
Review
The Mcm2-7 complex serves as the eukaryotic replicative helicase, the molecular motor that both unwinds duplex DNA and powers fork progression during DNA replication. Consistent with its central role in this process, much prior work has illustrated that Mcm2-7 loading and activation are landmark events in the regulation of DNA replication. Unlike any other hexameric helicase, Mcm2-7 is composed of six unique and essential subunits. Although the unusual oligomeric nature of this complex has long hampered biochemical investigations, recent advances with both the eukaryotic as well as the simpler archaeal Mcm complexes provide mechanistic insight into their function. In contrast to better-studied homohexameric helicases, evidence suggests that the six Mcm2-7 complex ATPase active sites are functionally distinct and are likely specialized to accommodate the regulatory constraints of the eukaryotic process.
Topics: Amino Acid Sequence; Animals; Cell Cycle Proteins; DNA; DNA Helicases; DNA Replication; DNA-Binding Proteins; Humans; Molecular Sequence Data; Multienzyme Complexes
PubMed: 19946136
DOI: 10.1128/MMBR.00019-09 -
Journal of Bacteriology Oct 2003
Review
Topics: Bacteria, Anaerobic; Cellulose; Gene Expression Regulation, Bacterial; Multienzyme Complexes; Temperature
PubMed: 14526000
DOI: 10.1128/JB.185.20.5907-5914.2003