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A new enzyme superfamily - The phosphopantetheinyl transferases
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Lambalot RH, Gehring AM, Flugel RS, Zuber P, LaCelle M, Marahiel MA, Reid R, Khosla C, Walsh CT A new enzyme superfamily - the phosphopantetheinyl transferases. Chem Biol. 3:1996;923-936.
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0024437776
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Gramicidin S biosynthesis operon containing the structural genes grsA and grsB has an open reading frame encoding a protein homologous to fatty acid thioesterases
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Krätzschmar J, Krause M, Marahiel MA Gramicidin S biosynthesis operon containing the structural genes grsA and grsB has an open reading frame encoding a protein homologous to fatty acid thioesterases. J Bacteriol. 171:1989;5422-5429.
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Penicillin biosynthesis: Energy requirement for tripeptide precursor formation by Δ-(L-β-aminoadipyl)-L-cysteinyl-D-valine synthetase from Acremonium chrysogenum
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Kallow W, von Döhren H, Kleinkauf H Penicillin biosynthesis: energy requirement for tripeptide precursor formation by Δ-(L-β-aminoadipyl)-L-cysteinyl-D-valine synthetase from Acremonium chrysogenum. Biochemistry. 37:1998;5947-5952.
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Kallow, W.1
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Kleinkauf, H.3
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0032562142
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Reconstitution and characterization of the Escherichia coli enterobactin synthetase from EntB, EntE, and EntF
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This paper describes the in vitro reconstitution of the biosynthesis of the siderophore enterobactin, a cyclic trilactone of N-(2,3-dihydroxybenzoyl)-L-serine. The necessary components were holo-EntB (isochorismate lyase-ArCP), EntE (dihydroxybenzoate-AMP ligase), and holo-EntF (C-A-PCP-TE synthetase). The holo enzymes were produced by the covalent attachment of a P-pant group catalyzed by EntD, which is a phosphopantetheinyl transferase. Mutation of the active site serine in the TE of EntF abolished enterobactin synthetase activity
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Gehring AM, Mori I, Walsh CT Reconstitution and characterization of the Escherichia coli enterobactin synthetase from EntB, EntE, and EntF. Biochemistry. 37:1998;2648-2659. This paper describes the in vitro reconstitution of the biosynthesis of the siderophore enterobactin, a cyclic trilactone of N-(2,3-dihydroxybenzoyl)-L-serine. The necessary components were holo-EntB (isochorismate lyase-ArCP), EntE (dihydroxybenzoate-AMP ligase), and holo-EntF (C-A-PCP-TE synthetase). The holo enzymes were produced by the covalent attachment of a P-pant group catalyzed by EntD, which is a phosphopantetheinyl transferase. Mutation of the active site serine in the TE of EntF abolished enterobactin synthetase activity.
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Gehring, A.M.1
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Walsh, C.T.3
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0032193282
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Iron acquisition in plague: Modular logic in enzymatic biogenesis of yersiniabactin by Yersinia pestis
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Gehring AM, DeMoll E, Fetherston JD, Mori I, Mayhew GF, Blattner FR, Walsh CT, Perry RD Iron acquisition in plague: modular logic in enzymatic biogenesis of yersiniabactin by Yersinia pestis. Chem Biol. 5:1998;573-586.
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Blattner, F.R.6
Walsh, C.T.7
Perry, R.D.8
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14
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0032211974
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Identification of a Mycobacterium tuberculosis gene cluster encoding the biosynthetic enzymes for assembly of the virulence-conferring siderophore mycobactin
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Quadri LEN, Sello J, Keating TA, Weinreb PH, Walsh CT Identification of a Mycobacterium tuberculosis gene cluster encoding the biosynthetic enzymes for assembly of the virulence-conferring siderophore mycobactin. Chem Biol. 5:1998;631-645.
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Quadri, L.E.N.1
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15
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0033617184
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Molecular characterization of the genes of actinomycin synthetase I and of a 4-methyl-3-hydroxyanthranilic acid carrier protein involved in the assembly of the acylpeptide chain of actinomycin in Streptomyces
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Pfennig F, Schauwecker F, Keller U Molecular characterization of the genes of actinomycin synthetase I and of a 4-methyl-3-hydroxyanthranilic acid carrier protein involved in the assembly of the acylpeptide chain of actinomycin in Streptomyces. J Biol Chem. 274:1999;12508-12516.
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J Biol Chem
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Pfennig, F.1
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Keller, U.3
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16
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0031736408
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Dihydroaeruginoic acid synthetase and pyochelin synthetase, products of the pchEF genes, are induced by extracellular pyochelin in Pseudomonas aeruginosa
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Reimmann C, Serino L, Beyeler M, Haas D Dihydroaeruginoic acid synthetase and pyochelin synthetase, products of the pchEF genes, are induced by extracellular pyochelin in Pseudomonas aeruginosa. Microbiology. 144:1998;3135-3148.
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Reimmann, C.1
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17
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0033574774
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Aminoacyl-CoAs as probes of condensation domain selectivity in nonribosomal peptide synthesis
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This paper describes a method for bypassing the amino acid substrate specificity of the adenylation domains in NRPSs. After chemically synthesizing aminoacylated-Coenzyme A analogs, a PPTase can be used to directly modify an apo-xCP domain with the P-pant tether carrying the desired monomer. These monomers can then test the specificity of the condensation domain toward the upstream and downstream substrates. The authors found much greater selectivity for the the downstream monomer than for the upstream one
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Belshaw PJ, Walsh CT, Stachelhaus T Aminoacyl-CoAs as probes of condensation domain selectivity in nonribosomal peptide synthesis. Science. 284:1999;486-489. This paper describes a method for bypassing the amino acid substrate specificity of the adenylation domains in NRPSs. After chemically synthesizing aminoacylated-Coenzyme A analogs, a PPTase can be used to directly modify an apo-xCP domain with the P-pant tether carrying the desired monomer. These monomers can then test the specificity of the condensation domain toward the upstream and downstream substrates. The authors found much greater selectivity for the the downstream monomer than for the upstream one.
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Science
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Belshaw, P.J.1
Walsh, C.T.2
Stachelhaus, T.3
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18
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0032506939
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Primer unit specificity in rifamycin biosynthesis principally resides in the later stages of the biosynthetic pathway
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Hunziker D, Yu T-W, Hutchinson CR, Floss HG, Khosla C Primer unit specificity in rifamycin biosynthesis principally resides in the later stages of the biosynthetic pathway. J Am Chem Soc. 120:1998;1092-1093.
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J Am Chem Soc
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Hunziker, D.1
Yu, T.-W.2
Hutchinson, C.R.3
Floss, H.G.4
Khosla, C.5
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19
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0030756031
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Structural basis for the activation of phenylalanine in the non-ribosomal biosynthesis of gramicidin S
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Conti E, Stachelhaus T, Marahiel MA, Brick P Structural basis for the activation of phenylalanine in the non-ribosomal biosynthesis of gramicidin S. EMBO J. 16:1997;4174-4183.
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Conti, E.1
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Brick, P.4
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0033179468
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The specificity-conferring code of adenylation domains in nonribosomal peptide synthetases
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Stachelhaus T, Mootz HD, Marahiel MA The specificity-conferring code of adenylation domains in nonribosomal peptide synthetases. Chem Biol. 6:1999;493-505.
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Stachelhaus, T.1
Mootz, H.D.2
Marahiel, M.A.3
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21
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0033514424
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Dissecting the role of acyltransferase domains of modular polyketide synthases in the choice and stereochemical fate of extender units
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Lau J, Fu H, Cane DE, Khosla C Dissecting the role of acyltransferase domains of modular polyketide synthases in the choice and stereochemical fate of extender units. Biochemistry. 38:1999;1643-1651.
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Lau, J.1
Fu, H.2
Cane, D.E.3
Khosla, C.4
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22
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0032575648
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Peptide bond formation in nonribosomal peptide biosynthesis
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This paper describes an assay for condensation domain function based on dipeptide formation and release by two modules: GrsA (PheATE) from gramicidin S synthetase, and truncated TycB (ProCAT) from tyrocidin synthetase. A functional condensation domain catalyzes amide bond formation, with noncatalytic release of a diketopiperazine molecule providing an assayable product. A mutation in the proposed active site motif of the condensation domain abolished the condensation reaction
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Stachelhaus T, Mootz HD, Bergendahl V, Marahiel MA Peptide bond formation in nonribosomal peptide biosynthesis. J Biol Chem. 273:1998;22773-22781. This paper describes an assay for condensation domain function based on dipeptide formation and release by two modules: GrsA (PheATE) from gramicidin S synthetase, and truncated TycB (ProCAT) from tyrocidin synthetase. A functional condensation domain catalyzes amide bond formation, with noncatalytic release of a diketopiperazine molecule providing an assayable product. A mutation in the proposed active site motif of the condensation domain abolished the condensation reaction.
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J Biol Chem
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Stachelhaus, T.1
Mootz, H.D.2
Bergendahl, V.3
Marahiel, M.A.4
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23
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0033574768
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Dissecting and exploiting intermodular communication in polyketide synthases
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The authors demonstrate that the individual modules of the DEBS PKS are tolerant toward the identity of the upstream acyl chain in each condensation cycle. They also describe linkers, short sequences that lie within and between the individual polypeptides, that are critical for retaining function when modules are swapped out of their original position. Proper engineering of linkers allows transfer of intermediates between unnaturally linked modules, and thus the catalytic production of 'unnatural natural products'
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Gokhale RS, Tsuji SY, Cane DE, Khosla C Dissecting and exploiting intermodular communication in polyketide synthases. Science. 284:1999;482-485. The authors demonstrate that the individual modules of the DEBS PKS are tolerant toward the identity of the upstream acyl chain in each condensation cycle. They also describe linkers, short sequences that lie within and between the individual polypeptides, that are critical for retaining function when modules are swapped out of their original position. Proper engineering of linkers allows transfer of intermediates between unnaturally linked modules, and thus the catalytic production of 'unnatural natural products'.
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Science
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Gokhale, R.S.1
Tsuji, S.Y.2
Cane, D.E.3
Khosla, C.4
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24
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0033529935
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Direct evidence that the rifamycin polyketide synthase assembles polyketide chains processively
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The antibiotic rifamycin is an undecaketide assembled by the rifA-rifE genes, and cyclized and released by the separate amide synthase encoded by rifF. Inactivation of this thioesterase abolishes rifamycin production, and instead leads to the accumulation of covalently attached linear polyketides at each carrier protein waystation, thus indicating that rifamycin synthase can simultaneously process multiple acyl chains
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Yu T-w, Shen Y, Doi-Katayama Y, Tang L, Park C, Moore BS, Hutchinson CR, Floss HG Direct evidence that the rifamycin polyketide synthase assembles polyketide chains processively. Proc Natl Acad Sci USA. 96:1999;9051-9056. The antibiotic rifamycin is an undecaketide assembled by the rifA-rifE genes, and cyclized and released by the separate amide synthase encoded by rifF. Inactivation of this thioesterase abolishes rifamycin production, and instead leads to the accumulation of covalently attached linear polyketides at each carrier protein waystation, thus indicating that rifamycin synthase can simultaneously process multiple acyl chains.
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Proc Natl Acad Sci USA
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Yu, T.-W.1
Shen, Y.2
Doi-Katayama, Y.3
Tang, L.4
Park, C.5
Moore, B.S.6
Hutchinson, C.R.7
Floss, H.G.8
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25
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0031459595
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The bacitracin biosynthesis operon of Bacillus licheniformis ATCC 10716: Molecular characterization of three multi-modular peptide synthetases
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Konz D, Klens A, Schörgendorfer K, Marahiel MA The bacitracin biosynthesis operon of Bacillus licheniformis ATCC 10716: molecular characterization of three multi-modular peptide synthetases. Chem Biol. 4:1997;927-937.
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Konz, D.1
Klens, A.2
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Marahiel, M.A.4
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26
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0032544193
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The nonribosomal peptide synthetase HMWP2 forms a thiazoline ring during biogenesis of yersiniabactin, an iron-chelating virulence factor of Yersinia pestis
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This paper describes the first biochemical characterization of a cyclization domain, the first of three in yersiniabactin synthetase. The amino-terminal two-thirds of the synthetase HMWP2 is able to catalyze bond formation between salicylate and cysteine, and then to cyclize and dehydrate this intermediate to a hydroxyphenyl-thiazoline-carboxylate. This product is released from the enzyme fragment by hydrolysis or by attack of cysteine
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Gehring AM, Mori I, Perry RD, Walsh CT The nonribosomal peptide synthetase HMWP2 forms a thiazoline ring during biogenesis of yersiniabactin, an iron-chelating virulence factor of Yersinia pestis. Biochemistry. 37:1998;11637-11650. This paper describes the first biochemical characterization of a cyclization domain, the first of three in yersiniabactin synthetase. The amino-terminal two-thirds of the synthetase HMWP2 is able to catalyze bond formation between salicylate and cysteine, and then to cyclize and dehydrate this intermediate to a hydroxyphenyl-thiazoline-carboxylate. This product is released from the enzyme fragment by hydrolysis or by attack of cysteine.
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Biochemistry
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Gehring, A.M.1
Mori, I.2
Perry, R.D.3
Walsh, C.T.4
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27
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0032830741
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Bleomycin biosynthesis in Streptomyces verticillus ATCC15003: A model of hybrid peptide and polyketide biosynthesis
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Shen B, Du L, Sanchez C, Chen M, Edwards DJ Bleomycin biosynthesis in Streptomyces verticillus ATCC15003: a model of hybrid peptide and polyketide biosynthesis. Bioorg Chem. 27:1999;155-171.
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Shen, B.1
Du, L.2
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Chen, M.4
Edwards, D.J.5
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28
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0033515090
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Multiple genetic modifications of the erythromycin polyketide synthase to produce a library of novel 'unnatural' natural products
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Beginning with the DEBS PKS system, the authors have described manipulation of several elements that enable combinatorial biosynthesis of the 6-deoxyerythronolide B (6-dEB) core structure - AT substitution (affecting monomer identity), KR deletion and addition (affecting the oxidation state of a carbon center), and KR stereochemical alteration (affecting sterochemistry of a carbon center). By making single, double, and triple substitutions in the PKS, over 100 novel 6-dEB analogs have been produced, although the titers of the new products were uneven. The authors emphasize the inherent 'plasticity' of the PKS, as demonstrated by their successful modifications of each of the six modules of DEBS
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McDaniel R, Thamchaipenet A, Gustafsson C, Fu H, Betlach M, Ashley G Multiple genetic modifications of the erythromycin polyketide synthase to produce a library of novel 'unnatural' natural products. Proc Natl Acad Sci USA. 96:1999;1846-1851. Beginning with the DEBS PKS system, the authors have described manipulation of several elements that enable combinatorial biosynthesis of the 6-deoxyerythronolide B (6-dEB) core structure - AT substitution (affecting monomer identity), KR deletion and addition (affecting the oxidation state of a carbon center), and KR stereochemical alteration (affecting sterochemistry of a carbon center). By making single, double, and triple substitutions in the PKS, over 100 novel 6-dEB analogs have been produced, although the titers of the new products were uneven. The authors emphasize the inherent 'plasticity' of the PKS, as demonstrated by their successful modifications of each of the six modules of DEBS.
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Proc Natl Acad Sci USA
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McDaniel, R.1
Thamchaipenet, A.2
Gustafsson, C.3
Fu, H.4
Betlach, M.5
Ashley, G.6
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Genetic evidence for a role of thioesterase domains, integrated in or associated with peptide synthetases, in non-ribosomal peptide biosynthesis in Bacillus subtilis
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Schneider A, Marahiel MA Genetic evidence for a role of thioesterase domains, integrated in or associated with peptide synthetases, in non-ribosomal peptide biosynthesis in Bacillus subtilis. Arch Microbiol. 169:1998;404-410.
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Manipulation of macrolide ring size by directed mutagenesis of a modular polyketide synthase
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Engineering of peptide synthetases. Key role of the thioesterase-like domain for efficient production of recombinant peptides
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De Ferra, F.1
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0029027352
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Repositioning of a domain in a modular polyketide synthase to promote specific chain cleavage
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Cortés J, Wiesmann KEH, Roberts GA, Brown MJB, Staunton J, Leadley PF Repositioning of a domain in a modular polyketide synthase to promote specific chain cleavage. Science. 268:1995;1487-1489.
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Cortés, J.1
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Impact of thioesterase activity on tylosin biosynthesis in Streptomyces fradiae
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Molecular and biochemical characterization of the protein template controlling biosynthesis of the lipopeptide lichenysin
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Konz D, Doekel S, Marahiel MA Molecular and biochemical characterization of the protein template controlling biosynthesis of the lipopeptide lichenysin. J Bacteriol. 181:1999;133-140.
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Xue Y, Zhao L, Liu H-w, Sherman DH A gene cluster for macrolide antibiotic biosynthesis in Streptomyces venezuelae: architecture of metabolic diversity. Proc Natl Acad Sci USA. 95:1998;12111-12116.
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0033150199
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Assembly line enzymology by multimodular nonribosomal peptide synthetases: The thioesterase domain of E. coli EntF catalyzes both elongation and cyclolactonization
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Through thioesterase mutagenesis and mass spectrometry of trapped intermediates, this paper confirms a dual role for the TE domain of enterobactin synthetase: it not only cyclolactonizes the trimer of N-(2,3-dihydroxybenzoate)-L-serine to yield enterobactin as the final step, but it also serves as the last waystation for elongation. In this way, enterobactin synthetase displays a novel architecture; for an NRPS constructing a product composed of six monomers, five modules corresponding to each condensation event would be expected. However, enterobactin synthetase has only one module, which requires the TE to accept three units of N-(2,3-dibydroxybenzoate)-L-serine before cyclolactonization and release
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Shaw-Reid CA, Kelleher NL, Losey HC, Gehring AM, Berg C, Walsh CT Assembly line enzymology by multimodular nonribosomal peptide synthetases: the thioesterase domain of E. coli EntF catalyzes both elongation and cyclolactonization. Chem Biol. 6:1999;385-400. Through thioesterase mutagenesis and mass spectrometry of trapped intermediates, this paper confirms a dual role for the TE domain of enterobactin synthetase: it not only cyclolactonizes the trimer of N-(2,3-dihydroxybenzoate)-L-serine to yield enterobactin as the final step, but it also serves as the last waystation for elongation. In this way, enterobactin synthetase displays a novel architecture; for an NRPS constructing a product composed of six monomers, five modules corresponding to each condensation event would be expected. However, enterobactin synthetase has only one module, which requires the TE to accept three units of N-(2,3-dibydroxybenzoate)-L-serine before cyclolactonization and release.
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Chem Biol
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Shaw-Reid, C.A.1
Kelleher, N.L.2
Losey, H.C.3
Gehring, A.M.4
Berg, C.5
Walsh, C.T.6
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Mechanism of alkaloid cyclopeptide synthesis in the ergot fungus Claviceps purpurea
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Walzel B, Riederer B, Keller U Mechanism of alkaloid cyclopeptide synthesis in the ergot fungus Claviceps purpurea. Chem Biol. 4:1997;223-230.
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Biosynthesis of ansatrienin (mycotrienin) and naphthomycin. Identification and analysis of two separate biosynthetic gene clusters in Streptomyces collinus Tü 1892
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Chen S, von Bamberg D, Hale V, Breuer M, Hardt B, Müller R, Floss HG, Reynolds KA, Leistner E Biosynthesis of ansatrienin (mycotrienin) and naphthomycin. Identification and analysis of two separate biosynthetic gene clusters in Streptomyces collinus Tü 1892. Eur J Biochem. 261:1999;98-107.
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Floss, H.G.7
Reynolds, K.A.8
Leistner, E.9
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Lysine biosynthesis in Saccharomyces cerevisiae: Mechanism of α-aminoadipate reductase (Lys2) involves posttranslational phosphopantetheinylation by Lys5
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Ehmann DE, Gehring AM, Walsh CT Lysine biosynthesis in Saccharomyces cerevisiae: mechanism of α-aminoadipate reductase (Lys2) involves posttranslational phosphopantetheinylation by Lys5. Biochemistry. 38:1999;6171-6177.
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Two multifunctional peptide synthetases and an O-methyltransferase are involved in the biosynthesis of the DNA-binding antibiotic and antitumour agent saframycin Mx1 from Myxococcus xanthus
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Pospiech A, Bietenhader J, Schupp T Two multifunctional peptide synthetases and an O-methyltransferase are involved in the biosynthesis of the DNA-binding antibiotic and antitumour agent saframycin Mx1 from Myxococcus xanthus. Microbiology. 142:1996;741-746.
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Characterization of the syringomycin synthetase gene cluster
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The authors describe the synthetase of syringomycin, a lipodepsipeptide with phytotoxic properties. Syringomycin synthetase is unique in its architecture in that the 'colinearity rule' describing the assembly line process is not followed. There are nine monomers incorporated into syringomycin; the adenylation domain for threonine, the last amino acid in the chain, is not located in the last module upstream of the TE domain, where it would be expected, but rather on a separate protein, SyrB, located upstream of the main SyrE polypeptide. This implies that, at least, a specific in trans threonyl-AMP transfer must occur from the SyrB protein to the last PCP in SyrE
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Guenzi E, Galli G, Grgurina I, Gross DC, Grandi G Characterization of the syringomycin synthetase gene cluster. J Biol Chem. 273:1998;32857-32863. The authors describe the synthetase of syringomycin, a lipodepsipeptide with phytotoxic properties. Syringomycin synthetase is unique in its architecture in that the 'colinearity rule' describing the assembly line process is not followed. There are nine monomers incorporated into syringomycin; the adenylation domain for threonine, the last amino acid in the chain, is not located in the last module upstream of the TE domain, where it would be expected, but rather on a separate protein, SyrB, located upstream of the main SyrE polypeptide. This implies that, at least, a specific in trans threonyl-AMP transfer must occur from the SyrB protein to the last PCP in SyrE.
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J Biol Chem
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Guenzi, E.1
Galli, G.2
Grgurina, I.3
Gross, D.C.4
Grandi, G.5
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