Identification of Novel Peptides Originated from Biodegradation of Microcystins Using UHPLC-HRMS/MS

Biodegradation of toxic cyanobacterial cyclic peptides microcystins (MCs) by heterotrophic bacteria isolated from freshwater has gained global interest as an eco-friendly approach for water treatment. This study focused on elucidating the structures of MC-LR breakdown products generated by a group of five bacteria previously isolated from Lake Erie. Three isomers of a tetrapeptide NH2-Adda-Glu-Mdha-Ala-OH (Adda-TetraP-1, 2, and 3) and a cyclic tripeptide NH2-Adda-Glu-Mdha-OH (Adda-TriP) were identified during the MC-LR degradation process. Of these peptides, only linear Adda-TetraP-1 was described previously. UHPLC-high-resolution (HR)-MS and MS/MS data acquired for tetrapeptides and the tripeptide and their thiol derivatization showed that Adda-TetraP-2, Adda-TetraP-3, and Adda-TriP were cyclic peptides that contained modified Mdha and Adda moieties. The experimental evidence suggests that the Adda amine group of Adda-TetraP-1 underwent intramolecular aza-Michael reaction with the Mdha double bond to form a cyclic tetrapeptide, which appeared in two diastereomeric forms, of which Adda-TetraP-2 was formed faster than Adda-Tetra-P-3. The Mdha-Ala peptide bond of Adda-TetraP-2 appeared to be hydrolyzed by bacteria to form Adda-TriP. Independent of our laboratory cultures, Adda-TetraP-1, 2, and 3 were detected in Lake Erie water during cyanobacterial blooms, and identifying them will assist in elucidating biodegradation pathways of MC congeners and genes relevant to MC degradation.