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Dorrestein P.C., Yeh E., Garneau-Tsodikova S., Kelleher N.L., and Walsh C.T. Dichlorination of a pyrrolyl-S-carrier protein by FADH2-dependent halogenase PltA during pyoluteorin biosynthesis. Proc Natl Acad Sci U S A 102 (2005) 13843-13848
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A Bottom Up MS assay using low-resolution instrumentation is presented as applied to the study of a prototypical PKS system, including the use of MS for competition and kinetic analysis of differential selection of a variety of intermediates by a ketosynthase domain. Active site peptides from multiple domains with covalently tethered intermediates were identified.
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The development and implementation of the Ppant ejection assay for NRPS and PKS analysis is discussed in detail, in addition to presentation of numerous examples of its applications in a variety of systems.
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Hansen D.B., Bumpus S.B., Aron Z.D., Kelleher N.L., and Walsh C.T. The loading module of mycosubtilin: an adenylation domain with fatty acid selectivity. J Am Chem Soc 129 (2007) 6366-6367
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Incorporation of nonmethyl branches by isoprenoid-like logic: multiple beta-alkylation events in the biosynthesis of myxovirescin A1
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Calderone C.T., Iwig D.F., Dorrestein P.C., Kelleher N.L., and Walsh C.T. Incorporation of nonmethyl branches by isoprenoid-like logic: multiple beta-alkylation events in the biosynthesis of myxovirescin A1. Chem Biol 14 (2007) 835-846
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This paper discusses the use of FTMS for elucidation of the mechanism of β-branch incorporation by isoprenoid-like logic in the biosynthesis of bacillaene, highlighting the ability of FTMS to confirm the entire reaction sequence. A similar mechanism is reported for a different system in [35].
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Calderone C.T., Kowtoniuk W.E., Kelleher N.L., Walsh C.T., and Dorrestein P.C. Convergence of isoprene and polyketide biosynthetic machinery: isoprenyl-S-carrier proteins in the pksX pathway of Bacillus subtilis. Proc Natl Acad Sci U S A 103 (2006) 8977-8982. This paper discusses the use of FTMS for elucidation of the mechanism of β-branch incorporation by isoprenoid-like logic in the biosynthesis of bacillaene, highlighting the ability of FTMS to confirm the entire reaction sequence. A similar mechanism is reported for a different system in [35].
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Harnessing the chemical activation inherent to carrier protein-bound thioesters for the characterization of lipopeptide fatty acid tailoring enzymes
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This article discusses the use of on-line LC-MS and the Ppant ejection assay in characterization of three enzymes involved in the biosynthesis of calcium-dependent antibiotic; FTMS and Ppant ejection were applied to confirm phosphopantetheinylation of a putative thiolation domain and characterize an oxidase with intrinsic epoxidase activity that acts at covalently bound intermediates.
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