A Promiscuous Path to Novel Guanidines: Arginine-Modifying Enzymes as Key Synthetic Players

The study falls into two sections. The 1st Section describes an efficient enzyme-catalysed process for the chemoselective (m)ethylation of various guanidine-containing molecules, including arginine analogues, derivatives, and oligopeptides. The process relies on the promiscuous methyltransferase SznE (or an engineered variant thereof) to transfer a methyl or ethyl group, from S-adenosylmethionine (SAM) or S-adenosylethionine (SAE), respectively. The enzyme MtnN is included to counteract the inhibitory effect of the byproduct S-adenosylhomocysteine. The tandem catalytic system proved highly effective for >35 guanidine-containing substrates, achieving 80-100% conversion and demonstrating its utility for late-stage functionalisation of complex natural products and drugs (e.g., the pentapeptide opiorphin). These findings highlight SznE's potential for biocatalysis, engineering, and industrial applications. The 2nd Section demonstrates the biocatalytic versatility of three amidinotransferases (SxtG1-3), when used alone or with an engineered α-oxoamine synthase domain (SxtAAOS), for synthesising various nitrogen-rich 2-aminoimidazole derivatives. A wide range of α-aminoketones of arginine, arginine derivatives, and analogues were prepared either synthetically or by leveraging the promiscuous C-C bond forming capacity of an engineered SxtA-AOS variant. These α-aminoketones serve as substrates for SxtG1-3, in the presence of amidino donors, to produce corresponding α-guanidinylated ketone intermediates that cyclise into 2-aminoimidazole derivatives. Computational analyses suggest that (double) hydrogen bonding catalyses the final cyclisation. The structural diversity of the resulting 2-aminoimidazole alkaloids showcases the flexibility of the early enzymes involved in saxitoxin biosynthesis and the engineerability of the pathway for producing novel saxitoxins with therapeutic and mechanistic enzymology potential.

Grantor dc:publisher.institution

University of Bradford

Author dc:creator

• Fakhry, Ayman A.A.

Advisor dc:contributor.advisor

• Hamed, Refaat

• Methyltransferase
• Amidinotransferase
• PLP-dependent enzyme
• Oxamine synthase
• Biocatalysis
• Protein engineering
• Biosynthesis
• Enzymology
• Guanidines
• Arginine-modifying enzymes

Statement dc:rights

• <a rel="license" href="http://creativecommons.org/licenses/by-nc-nd/3.0/"><img alt="Creative Commons License" style="border-width:0" src="http://i.creativecommons.org/l/by-nc-nd/3.0/88x31.png" /></a><br />The University of Bradford theses are licenced under a <a rel="license" href="http://creativecommons.org/licenses/by-nc-nd/3.0/">Creative Commons Licence</a>.

Language dc:language.iso

en

Repository record dc:identifier.uri

https://bradscholars.brad.ac.uk/handle/10454/20839

OAI identifier oai:identifier

oai:bradscholars.brad.ac.uk:10454/20839