Application of a substrate-tolerant lanthipeptide synthetase in bioengineering peptide natural products

Lanthipeptides constitute a diverse class of peptide natural products defined by the presence of lanthionine residue(s). These peptides belong to a larger family of natural products called RiPPs, which stands for ribosomally synthesized and post-translationally modified peptides. Like many RiPPs classes, the enzymes involved in the biosynthesis of lanthipeptides demonstrate relatively high substrate tolerance, a feature which, combined with the ribosomal origin of the peptide substrates, facilitates bioengineering efforts of these peptides via evolution techniques. This dissertation is thematically built around an intriguing example of extreme catalytic promiscuity in lanthipeptides biosynthesis, the prochlorosin system. The journey starts with a discussion of why and how nature has possibly evolved the prochlorosins to favor diversity-oriented biosynthesis, followed by an overview of high-throughput screening methods that have been applied to lanthipeptides. In chapter 2, the biosynthesis of prochlorosins is shown to be under substrate control such that the site-selectivity of lanthionine formation is heavily influenced by the substrates’ primary sequences. This feature opens up an opportunity for the construction of lanthipeptide libraries which is realized in chapter 3, in which an in-liposome directed evolution experiment is described in order to discover lanthipeptides with improved bioactivity. Finally, in chapter 4, the prochlorosin biosynthetic machinery is combined with an enzyme from a distant RiPP class in order to create hybrid structures that have potentials for higher stability, affinity, or adaptability for high-throughput screening experiments.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Chemistry

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2023

Author dc:creator

• Le, Tung Thanh

Contributors dc:contributor

• van der Donk, Wilfred A.
• Silverman, Scott K.
• Mitchell, Douglas A.
• Zhao, Huimin

• Lanthipeptides
• Enzymatic Promiscuity
• Bio-engineering
• Ripps

Statement dc:rights

• Copyright 2023 Tung Le

Language dc:language

en, eng

Handle dc:identifier

https://hdl.handle.net/2142/121960

OAI record oai:identifier

https://www.ideals.illinois.edu/oai-pmh?verb=GetRecord&metadataPrefix=etdms&identifier=oai%3Awww.ideals.illinois.edu%3A2142%2F121960