Synthesis of the non-proteinogenic amino acid, (4S)-4,5-dihydroxy-L-norvaline, and its application in the total synthesis of an Euglenatide B analog

Melisa S. Gonzalez, Rongrong Yu, Xiang Li, Rebecca A. Butcher

University of Florida

Euglenatides A-E are hybrid polyketide-nonribosomal cyclic peptides that were isolated in 2022 from the unicellular microalga Euglena gracilis. These compounds exhibit inhibitory activity towards pathogenic fungi and cancer cell lines, which indicates their potential as pharmaceuticals. In particular, euglenatide B, which was isolated in highest yield, consists of a macrolactam ring containing two asparagine (Asn) residues and three non-proteinogenic amino acids, namely b-aminoisobutyric acid (b-Aib), (4S)-4,5-dihydroxy-L-norvaline (Dnv) and the polyketide derived b-amino-2,5-dihydroxy-7-methoxy-8,10,12-eicosatrienoic acid. Interestingly, euglenatides are the first peptide natural products reported to contain the unusual amino acid, Dnv. Intrigued by nature’s design and the significant antiproliferative activity of euglenatides, a less structurally complex analog of euglenatide B was designed for bioactivity studies. A retrosynthetic approach started with the division of the analog into three fragments, fragment A: a simpler polyketide derived amino acid, (3R,5S)-3-amino-5-hydroxyoctadecanoic acid; fragment B: an Fmoc-protected version of Dnv, fragment C: the tripeptide N-b-Aib-Asn-Asn-C. Fragment A was synthesized in seven steps starting from tetradecanal and employing reactions such as Maruoka’s asymmetric allylation, olefin oxidative cleavage, and asymmetric synthesis of tert­-butanesulfinyl-protected amines. Fragment B was synthesized in five steps employing O’Donnell’s asymmetric allylation and Sharpless asymmetric dihydroxylation reactions. Fragment C was produced using a solid-phase peptide synthesizer. Future directions will focus on testing antiproliferative activity of the synthesized analog against pathogenic fungi and human cancer cells. If the compound does shows bioactivity, the synthetic route will be modified to include a photoaffinity tag for protein pull down studies. Additionally, more complex analogs will be synthesized to assess their antiproliferative potential.