Covalent flavinylation of 6‐hydroxy‐D‐nicotine oxidase involves an energy‐requiring process

Brandsch, Roderich; Bichler, Veronika

doi:10.1016/0014-5793(87)80433-7

Cited by 26 publications

(18 citation statements)

References 8 publications

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“…Following hyperexpression of the 6HDNO gene in E. coli, more than 50% of the expressed enzyme is found in the apo-form (Brandsch & Bichler, 1987). The apoenzyme can be reconstituted using FAD and an "energy-generating" system, consisting of ATP, phosphoenolpyruvate, and pyruvate kinase.…”

Section: -Hydroxy-~-nicotine Oxidase (Su-n3-histidyl Fad)mentioning

confidence: 99%

“…How is it that C-terminal truncated proteins, with as much as 40% of the sequence missing, are able to fold into native, apo-enzymes with properly formed FAD-binding sites? Is it possible that the N-terminal and C-terminal portions fold independently into separate domains?Following hyperexpression of the 6HDNO gene in E. coli, more than 50% of the expressed enzyme is found in the apo-form (Brandsch & Bichler, 1987). The apoenzyme can be reconstituted using FAD and an "energy-generating" system, consisting of ATP, phosphoenolpyruvate, and pyruvate kinase.…”

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confidence: 99%

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Covalent attachment of flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) to enzymes: The current state of affairs

1998

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The first identified covalent flavoprotein, a component of mammalian succinate dehydrogenase, was reported 42 years ago. Since that time, more than 20 covalent flavoenzymes have been described, each possessing one of five modes of FAD or FMN linkage to protein. Despite the early identification of covalent flavoproteins, the mechanisms of covalent bond formation and the roles of the covalent links are only recently being appreciated. The main focus of this review is, therefore, one of mechanism and function, in addition to surveying the types of linkage observed and the methods employed for their identification. Case studies are presented for a variety of covalent flavoenzymes, from which general findings are beginning to emerge.Keywords: covalent flavoproteins; flavin adenine dinucleotide; flavin mononucleotide; flavinylation; redox cofactors Cofactors are recruited by protein molecules to extend the chemistry available within the active sites of enzymes. The chemistries displayed are variable and the range of protein-specific cofactors available bears testimony to the types of biological reaction achieved by many enzyme molecules. Cofactors may be present in the freely dissociable form, but others, such as lipoic acid and biotin, are permanently linked to the host protein. Other cofactors (e.g., heme and pyridoxyl phosphate) fall into both categories in that they can be attached covalently to the protein or operate in the freely dissociable form. The covalent addition of specific cofactors to unique residues within a polypeptide chain is a fundamental problem in studies of biological molecular recognition. In many cases, this specificity is purchased at the expense of recruiting modification enzymes that direct the covalent addition of a cofactor to the host protein. For example, the additions of biotin and lipoic acid to the €-amino groups of specific Lys residues in carboxyltransferases and the 2-oxoacid dehydrogenases are directed by a biotin holoenzyme synthase (Sweetman et al., 1985;Takai et al., 1987) and Reprint requests to: N.S. Scmtton, Department of Biochemistry, University of Leicester, Adrian Building, University Road, Leicester LE1 7RH UK; e-mail: nss4@le.ac.uk; or W.S. McIntire, Molecular Biology Division, Department of Veterans Affairs Medical Center, San Francisco, California 94121; e-mail: wsm@itsa.ucsf.edu.lipoate-protein ligases (Brookfield et al., 1991; Moms et al., 1994), respectively. In a similar fashion, a heme cytochrome c lyase (Steiner et al., 1996) is responsible for the covalent addition of heme to two Cys residues via thioether linkages. In some enzymes, an existing side chain is modified to yield what is in effect a covalently attached cofactor, although no pre-existing cofactor molecule is incorporated into the enzyme. Several examples of this type of modification are provided in the next paragraph.For histidine decarboxylase of Lactobacillus 30A (Recsei & Snell, 1984), a pyruvoyl group is generated from an existing residue, Ser 82. The enzyme is synthesized as a proenzym...

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Section: -Hydroxy-~-nicotine Oxidase (Su-n3-histidyl Fad)mentioning

confidence: 99%

mentioning

confidence: 99%

Covalent attachment of flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) to enzymes: The current state of affairs

1998

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“…[14C]Riboflavin, 13'S]Met and t3,P]ATP were purchased from Amersham Buchler, (Braunschweig, FRG) ; [14C]FAD was synthesized enzymatically [6]. Protein-A -Sepharose was from Pharmacia (Freiburg, FRG).…”

Section: Chemicalsmentioning

confidence: 99%

Rat liver dimethylglycine dehydrogenase

Lang

Polster

Brandsch

1991

European Journal of Biochemistry

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Dimethylglycine dehydrogenase (Me2GlyDH), an enzyme of choline catabolism specifically expressed in the mammalian liver, was analyzed in rat hepatocytes in culture. This mitochondrial enzyme carries the FAD cofactor covalently attached to the polypeptide chain by its riboflavin 8a position to N" of histidine [Cook, R., Misono, Biol. Chem. 256,4102-41081, and a 43-amino-acid leader peptide. The Nterminus of Me,GlyDH contains a conserved amino acid sequence which forms the dinucleotide-binding site in many enzymes with noncovalently bound FAD. Close to the modified histidine there is an amino acid sequence resembling a sequence conserved in thymidylate synthases and shown in these enzymes to be involved in the binding of the pteroyl polyglutamate cofactor.Choline, taken up with the food or set free from phosphatidylcholine, is degraded in mammals, specifically in the liver. The enzymes of this pathway are mitochondrially located. The first, choline dehydrogenase, situated on the matrix side of the inner mitochondrial membrane, contains FAD and has a function similar to the succinate dehydrogenase complex in relation to the respiratory chain [l]. Two additional flavoenzymes of the pathway, dimethylglycine dehydrogenase (Me,GlyDH) and sarcosine dehydrogenase, were shown to contain, besides tightly bound pteroyl pentaglutamate, FAD covalently attached to the polypeptide chain by its riboflavin 8a position to N" of histidine [2].We have investigated the biogenesis of this covalent modification in 6-hydroxy-~-nicotine oxidase, an enzyme of procaryotic origin. With this particular enzyme the histidyl His(FAD) linkage is synthesized in an autocatalytic process by the polypeptide itself. It is not known how this protein modification is accomplished in enzymes of the eucaryotic Correspondence to R. Brandsch, Biochemisches Institut, Universitat Freiburg, Hermann-Herder-Str. 7, W-7800 Freiburg, Federal Republic of Germany Abbreviations. His(FAD), FAD linked through riboflavin position 8r to N" of histidine; MeZGlyDH, dimethylglycine dehydrogenase.Enzyme. Dimethylglycine dehydrogenase (EC 1.5.99.2). Nore. The nucleotide sequence corresponding to the coding part of the sequence reported in this paper is available from the EMBL data bank with accession number X55995.cell. In the case of other cofactors, covalent attachment to the protein, like biotin in carboxylases or heme in cytochrome c, requires a special enzyme [3, 41. The attachment of the prosthetic group is a step involved in the import of apocytochrome c into the intermembrane space and in its correct location within the inner mitochondrial membrane [S]. Whether a similar mitochondrial factor is involved in the covalent attachment of FAD to flavoenzymes is not known. It would also be of interest to know at which stage in the pathway from the precursor protein to the mature form the covalent modification takes place and how it influences the folding of the polypeptide chain on its way to the mature, enzymatically active form.In order to address these questions, we have...

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“…ROS, as well as reactive nitrogen species, are important signaling molecules in cardiovascular cells. Among ROS are superoxide anions, hydroxyl radicals and H 2 O 2 , which all take part in growth and apoptosis, as well as the expression of proinflammatory compounds and the modulation of endothelial functions Brandsch and Bichler (1987); Cross and Jones (1986); Cross et al (1984); Gatley and Sherratt (1976);O'Donnell et al (1993O'Donnell et al ( , 1994; Stuehr et al (1991) AEBSF (non-peptide) Oxidase assembly inhibitor: inhibits association of Nox2 with p47phox.…”

Section: Nox Inhibitors In Hypertensionmentioning

confidence: 99%