Journal Article

PUBDB-2017-10921

http://join2-wiki.gsi.de/foswiki/pub/Main/Artwork/join2_logo100x88.png Total Biosynthesis of the Pyrrolo[4,2]benzodiazepine Scaffold Tomaymycin on an In Vitro Reconstituted NRPS System

von Tesmar, A. ; Hoffmann, M. ; Pippel, J. ; Fayad, A. A. ; Dausend-Werner, S. ; Bauer, A. ; Blankenfeldt, W.Extern* ; Müller, R. (Corresponding author)

2017
Elsevier Amsterdam

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Please use a persistent id in citations: doi:10.1016/j.chembiol.2017.08.001

Abstract: In vitro reconstitution and biochemical analysis of natural product biosynthetic pathways remains a challenging endeavor, especially if megaenzymes of the nonribosomal peptide synthetase (NRPS) type are involved. In theory, all biosynthetic steps may be deciphered using mass spectrometry (MS)-based analyses of both the carrier protein-coupled intermediates and the free intermediates. We here report the “total biosynthesis” of the pyrrolo[4,2]benzodiazepine scaffold tomaymycin using an in vitro reconstituted NRPS system. Proteoforms were analyzed by liquid chromatography (LC)-MS to decipher every step of the biosynthesis on its respective megasynthetase with up to 170 kDa in size. To the best of our knowledge, this is the first report of a comprehensive analysis of virtually all chemical steps involved in the biosynthesis of nonribosomally synthesized natural products. The study includes experiments to determine substrate specificities of the corresponding A-domains in competition assays by analyzing the adenylation step as well as the transfer to the respective carrier protein domain.

Note: online first

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Contributing Institute(s):

1. DOOR-User (DOOR)

Research Program(s):

1. 6G3 - PETRA III (POF3-622) (POF3-622)

Experiment(s):

1. PETRA Beamline P11 (PETRA III)

Appears in the scientific report 2017

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Database coverage:
; Ebsco Academic Search ; NCBI Molecular Biology Database

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