Genome mining and annotation approval Georges Cohen Institut Pasteur Paris e-mail:gncohen@pasteur.fr .


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Genome mining and annotation validation Georges Cohen Institut Pasteur Paris e-mail:gncohen@pasteur.fr. As many as 40% of all predicted genes in completed prokaryotic genomes have no functional annotation.
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Genome mining and comment approval Georges Cohen Institut Pasteur Paris e-mail:gncohen@pasteur.fr

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As numerous as 40% of all anticipated qualities in finished prokaryotic genomes have no utilitarian explanation

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Many qualities have an anticipated capacity, yet that expectation has not been tentatively approved

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As numerous as 5-10% of anticipated quality capacities might be off base

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Many known compounds have no comparing qualities recognized in the succession databases

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Lysine aging - Known since the 50\'s - 1 mole of lysine is debased to 1 mole of acetate,1 mole of butyrate and 2 moles NH3 Well concentrated on in Clostridium sticklandii , additionally introduce in Porphyromonas gingivalis and Fusobacterium nucleatum

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Acyl-CoA dehydrogenase Not sequenced Butyrate-CoA acetoacetyl-CoA transférase Acetyl-CoA acetyltransférase Lysine maturation in Fusobacterium nucleatum Lysine 2,3-aminomutase N H N C O H 2 b - Lysine 5,6-aminomutase 2 H N C O H 2 kamA N H 2 kamD,E L y s i n e N H N H 2 C O H Not sequenced Not sequenced AtoA,D

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Best competitor: FN1867 Data digging for the 3,5-diaminohexanoate dehydrogenase encoding quality H 2 O + NADH + H + NAD + NH 3 Characteristics of 3,5-diaminohexanoate dehydrogenase: - disengaged and cleaned from Clostridium SB4, Clostridium sticklandii, Brevibacterium L5 - cofactor: NAD+ - atomic weight somewhere around 37 and 39 kDa - dimer or tetramer  Search for a F.nucleatum protein which a) has a coupling site for NAD+ b) has a sub-atomic weight around 38 kDa

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* Substrate  L-erythro-3,5-diaminohexanoate 2 stereoisomeric focuses  4 stereoisomers L-erythro D-erythro L-threo D-threo

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Synthesis of DL-erythro-3,5-diaminohexanoate Références: Chem. Berichte 1904, 37 , 2357-2362 Organic Preparations and Procedures Int. 1973, 5 , 31-35 + NH 3 + HCl 6 h reflux 150 °C, 20 h Under weight Sorbic corrosive DL-erythro-3,5-diaminohexanoate - Separation of erythro and threo by recrystallisation in isopropanol - no partition of the D et L isomers

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Acyl-CoA dehydrogenase Not sequenced Butyrate-CoA acetoacetyl-CoA transférase Acetyl-CoA acetyltransférase Lysine maturation in Fusobacterium nucleatum Lysine 2,3-aminomutase N H N C O H 2 b - Lysine 5,6-aminomutase 2 H N C O H 2 kamA N H 2 kamD,E L y s i n e N H N H 2 C O H Not sequenced Not sequenced AtoA,D

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1) Let the result of FN1867 aggregate FN1867 N H N H + 2 NAD NH 4 + H 2 O NADH H + C O H L-erythro - 3,5-DAH 3-Keto-5-aminohexanoate 2) Add then FN1868 and the co-substrate acetyl CoA FN1868 + acetyl-CoA N H O 2 3-Keto-5-aminohexanoate 3-aminobutyryl-CoA S C o A N H O 2 C O H O 3) Follow the vanishing of\'acetyl CoA utilizing citrate synthase(CS) O H CS acetyl-CoA + oxaloacetate+ DTNB citrate + CoA-disulfite + thionitrobenzoate absorbance at 412 nm Enzymatic test for FN 1868

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Tri-coupled examine for FN1869 FN1867 FN1868 Diaminohexanoate - > 3-keto-5-aminohexanoate - >3-aminobutyryl CoA FN1869 - > Crotonyl CoA

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Annett Kreimeyer Alain Perret Claudine Médigue Marcel Salanoubat Jean Weissenbach J.Biol.Chem.,(2007)282,7191-7 Georges Cohen, specialist

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