Exploring modular reengineering strategies to redesign the teicoplanin non-ribosomal peptide synthetase

Kaniušaitė, Milda; Goode, Robert J. A.; Tailhades, Julien; Schittenhelm, Ralf B.; Cryle, Max J.

doi:10.1039/d0sc03483e

Cited by 24 publications

(27 citation statements)

References 52 publications

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“…Apart from being unfavourable when it comes to bio‐synthetic production of specific bioactive compounds for pharmaceutical application, this finding suggests that it is possible to repurpose elongation modules to initiate peptide biosynthesis—at least under certain conditions—as most recently reported in an in vitro study. [45] …”

Section: Resultsmentioning

confidence: 99%

Synthetic Zippers as an Enabling Tool for Engineering of Non‐Ribosomal Peptide Synthetases**

et al. 2021

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Non-ribosomal peptide synthetases (NRPSs) are the origin of aw ide range of natural products,i ncluding many clinically used drugs.Efficient engineering of these often giant biosynthetic machineries to produce novel non-ribosomal peptides (NRPs) is an ongoing challenge.H ere we describe ac loning and co-expression strategy to functionally combine NRPS fragments of Gram-negative and -positive origin, synthesising novel peptides at titres up to 220 mg L À1 .E xtending from the recently introduced definition of eXchange Units (XUs), we inserted synthetic zippers (SZs) to split single protein NRPSs into independently expressed and translated polypeptide chains.T hese synthetic type of NRPS (type S) enables easier access to engineering,o vercomes cloning limitations,a nd provides as imple and rapid approach to building peptide libraries via the combination of different NRPS subunits.

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Section: Resultsmentioning

confidence: 99%

Synthetic Zippers as an Enabling Tool for Engineering of Non‐Ribosomal Peptide Synthetases**

et al. 2021

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“…Whilst the modular architecture of NRPSs has attracted great interest from the perspective of biosynthetic engineering, [17][18][19] such efforts have not always been successful. This can be attributed to the complexity of the machinery combined with the necessity for non-native substrates to pass through multiple catalytic domains, each of which imparts a degree of speci city.…”

Section: Introductionmentioning

confidence: 99%

Understanding condensation domain selectivity in non-ribosomal peptide biosynthesis: structural characterization of the acceptor bound state

Cryle¹,

Izoré²,

Ho³

et al. 2021

Preprint

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Non-ribosomal peptide synthetases are important enzymes for the assembly of complex peptide natural products. Within these multi-modular assembly lines, condensation domains perform the central function of chain assembly, typically by forming a peptide bond between two peptidyl carrier protein (PCP)-bound substrates. In this work, we report the first structural snapshots of a condensation domain in complex with an aminoacyl-PCP acceptor substrate. These structures allow the identification of a mechanism that controls access of acceptor substrates to the active site in condensation domains. The structures of this previously uncharacterized complex also allow us to demonstrate that condensation domain active sites do not contain a distinct pocket to select the side chain of the acceptor substrate during peptide assembly but that residues within the active site motif can instead serve to tune the selectivity of these central biosynthetic domains.

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“…Whilst the modular architecture of NRPSs has attracted great interest from the perspective of biosynthetic engineering [17][18][19] , such efforts have not always been successful. This can be attributed to the complexity of the machinery combined with the necessity for non-native substrates to pass through multiple catalytic domains, each of which imparts a degree of specificity.…”

mentioning

confidence: 99%

Structures of a non-ribosomal peptide synthetase condensation domain suggest the basis of substrate selectivity

Izoré

Kaczmarski

et al. 2021

Nat Commun

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Non-ribosomal peptide synthetases are important enzymes for the assembly of complex peptide natural products. Within these multi-modular assembly lines, condensation domains perform the central function of chain assembly, typically by forming a peptide bond between two peptidyl carrier protein (PCP)-bound substrates. In this work, we report structural snapshots of a condensation domain in complex with an aminoacyl-PCP acceptor substrate. These structures allow the identification of a mechanism that controls access of acceptor substrates to the active site in condensation domains. The structures of this complex also allow us to demonstrate that condensation domain active sites do not contain a distinct pocket to select the side chain of the acceptor substrate during peptide assembly but that residues within the active site motif can instead serve to tune the selectivity of these central biosynthetic domains.

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Exploring modular reengineering strategies to redesign the teicoplanin non-ribosomal peptide synthetase

Abstract: Non-ribosomal peptide synthesis is an important biosynthesis pathway in secondary metabolism. In this study we have investigated modularisation and redesign strategies for the glycopeptide antibiotic teicoplanin. Using the relocation or...

Cited by 24 publications

References 52 publications

Synthetic Zippers as an Enabling Tool for Engineering of Non‐Ribosomal Peptide Synthetases**

Synthetic Zippers as an Enabling Tool for Engineering of Non‐Ribosomal Peptide Synthetases**

Understanding condensation domain selectivity in non-ribosomal peptide biosynthesis: structural characterization of the acceptor bound state

Structures of a non-ribosomal peptide synthetase condensation domain suggest the basis of substrate selectivity

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