Mechanistic and synthetic studies on the prochlorosin and cytolysin families of lanthipeptides

Peptides are an attractive class of therapeutics, occupying a niche between small molecules and biologics. Research in the van der Donk lab focuses on lanthipeptides, a class of ribosomally synthesized and post-translationally modified peptides (RiPPs) that commonly feature antibacterial activity and contain the characteristic thioether residues lanthionine (Lan) and methyllanthionine (MeLan). Installation of thioether crosslinks in lanthipeptide biosynthesis is carried out by designated synthetases and involves dehydration of Ser/Thr residues and cyclization via Michael-type addition. The remarkably broad substrate scope of the synthetase ProcM inspired us to explore its mechanism in detail (chapter 2). My studies on ProcM revealed the directionality of dehydration, the order of cyclization, and that, despite the impressive substrate scope, none of the cyclizations are non-enzymatic. In collaboration, we established the irreversibility of the Michael-type addition and proposed that the topology of the thioether rings is under kinetic control. Solid phase peptide synthesis (SPPS) was used to generate the substrates to study ProcM, and is also a flexibile tool to access non-native lanthipeptide analogues. Interestingly, a lanthipeptide, cytolysin S (CylLS”), exhibited cytolytic activity in synergy with cytolysin L (CylLL”). Given that a thioether crosslink in CylLS” has an unusual LL-stereochemistry, the synthesis of a diastereomer of CylLS” with the more common DL-stereochemistry was achieved by SPPS (chapter 3). We probed whether the cytolytic activity depended on the LL-stereochemistry observed in CylLS”. Surprisingly, the unusual LL-stereochemistry was found to be important for the antibacterial activity, but not necessary for the hemolytic activity of CylLS”. I have also synthesized another hydrophobic lanthipeptide analogue, the portion of microbisporicin that contains the A and B ring (chapter 4). We established that this motif is not recognized by the halogenase MibH, and that the C terminus of microbisporicin is necessary for the chlorination by MibH. During my graduate studies, I had the opportunity to collaborate in a different area of research in our laboratory, the phosphonates. My efforts in the syntheses of various substrates and intermediates were instrumental in elucidating the biosynthetic pathways of dehydrophos, fosfazinomycin, and rhizocticin (chapter 5).

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

2016

Author dc:creator

• Mukherjee, Subha

Contributors dc:contributor

• van der Donk, Wilfred Adrianus
• Burke, Martin D.
• Metcalf, William W.
• Zhao, Huimin

• ribosomally synthesized and post-translationally modified peptides (RiPPs)
• Lanthipeptide
• Lanthionine
• antibiotics
• antimicrobial resistance
• prochlorosin
• ProcM
• Solid phase peptide synthesis (SPPS)
• lanthipeptide synthesis
• hydrophobic peptide synthesis
• dehydroalanine
• Michael-type addition
• non-enzymatic cyclization
• cytolysin
• microbisporicin
• phosphonate
• dehydrophos
• fosfazinomycin
• rhizocticin

Statement dc:rights

• Copyright 2015 Subha Mukherjee

Language dc:language

en

Handle dc:identifier

http://hdl.handle.net/2142/89174

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/89174