Novel heterocyclic peptides as potential inhibitors of proteases

In the 1970s, Clarke and Lowe modified papain with phenacylbromides to convert reactive cysteine 25 to serine 25 as a way to study papain binding. When the phenacylsulfide derivative of papain is exposed to UV light, it can undergo a Norrish Type II cleavage. Expanding on the work of Clarke and Lowe, the Myers group demonstrated cysteines in short peptides can be modified using this method to generate several species including heterocycles. One product of the Norrish Type II reaction is a thioaldehyde that subsequently tautomerizes and deprotonates to the enethiolate, which can be oxidized to the isothiazolone. Isothiazolones are of interest because they are already used as biocides and fungicides in agriculture and may have potential uses as inhibitors of cysteine proteases implicated in a wide variety of diseases. If the isothiazolone moiety can be introduced to a peptide or peptide mimic, it may be recognised by a cysteine protease and cause inhibition. Currently, one of the most popular methods for producing short peptides (>20 residues) is solid phase peptide synthesis (SPPS). SPPS can include additional steps to modify the peptides when they are still attached to the resin, which makes removal of waste and purification easier. The main goal of this project is to introduce the isothiazolone moiety with the peptide still attached to the SPPS resin. SPPS was used in this project to synthesize the tripeptide AlaCysAla as a model tripeptide which was modified on-resin. The most common protecting group for cysteine, the trityl group, was replaced with the mono-methoxytrityl (Mmt) group to allow for cysteine deprotection and modification without peptide cleavage. Initially, AlaCysAla was modified on-resin using methods from in solution cysteine modification and subsequently new methods of modification more optimal for on-resin cysteine modification were explored with the goal of producing isothiazolones in high yield and efficiency (Figure 1). During these optimisation experiments isothiazolone and another heterocycle, thiazole, were introduced to the peptide on-resin but isothiazolone was not produced in high yield. The successful introduction of these heterocycles to the tripeptide on-resin is a promising start and with more optimisation and understanding of the reactions involved, it should be possible to make isothiazolone containing peptides on-resin with high efficiency.

Funder

Science Foundation Ireland

Publisher

University of Galway

Collections

University of Galway Theses (Research Masters Theses)