Načrtovanje in sinteza N-aciliranih derivatov glukozamina kot zaviralcev MurA

Bezjak, Alen (author), Frlan, Rok (mentor)

Odpornost proti antibiotikom v bakterijah postaja vse bolj resen problem za globalni zdravstveni sistem, saj postajajo številne prej ozdravljive okužbe manj ozdravljive ali celo neozdravljive. Svetovna zdravstvena organizacija (WHO) je bakterijsko odpornost proti antibiotikom uvrstila med tri največje grožnje za javno zdravstvo v 21. stoletju. Razvoj novih protimikrobnih učinkovin tako postaja ena glavnih prioritet, saj se lahko le na ta način izognemo tej težavi. Peptidoglikan je pomemben gradnik bakterijske celične stene, ki se sintetizira v treh različnih delih celice: v citoplazmi, celični membrani in zunaj celice. Prva stopnja reakcijske poti poteka v citoplazmi, kjer encim MurA katalizira pomemben korak. Večina trenutnih protibakterijskih zdravil zavira sintezo peptidoglikana v zadnjih stopnjah sinteze, ki potekajo izven celice. Kljub temu nekaj antibiotikov deluje znotraj celice, med njimi je fosfomicin, ki zavira aktivnost encima MurA v citoplazmi. Ta magistrska naloga se osredotoča na sintezo dveh derivatov glukozamina, ki temeljita na molekularnem modeliranju in posnemanju naravnega substrata. Za pravilno usmerjenost funkcionalnih skupin in sintezo reverzibilnih zaviralcev smo kot osnovno spojino uporabili N-acetilglukozamin, ki lahko prodira skozi celično steno in membrano v notranjost bakterij, kjer se nahaja encim MurA. Glavni cilj je bil razviti sintezno pot za pripravo spojin, v katerih so kisli fragmenti pripeti na hidroksilno skupino na mestu 6. Najprej smo zaščitili vse funkcionalne skupine N-acetilglukozamina, razen hidroksilne skupine na mestu 6. Spojino 11 smo sintetizirali z direktno uvedbo ocetne kisline na hidroksilno skupino, s pomočjo kloroocetne kisline. Pri sintezi spojine 9 pa smo sprva morali hidroksilno skupino aktivirati s tvorbo tozilata, preko katerega smo potem tvorili derivat z dimetil malonatom, ki smo ga na koncu hidrolizirali do karboksilne kisline. Z encimskim testom smo potrdili, da sta obe spojini zavirali encimsko aktivnost MurA, pri čemer je imela spojina 9 vrednost IC50 84 μM in spojina 11 vrednost IC50 14 µM.

• Language: Slovenian
• Keywords: odpornost proti antibiotikom, peptidoglikan, zaviralec, N-acetilglukozamin, encim MurA
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2023
• PID: 20.500.12556/RUL-145886
• Publication date in RUL: 17.05.2023

Secondary language

• Language: English
• Title: Design and synthesis of N-acylated glucosamine derivatives as MurA inhibitors

Abstract

Antibiotic resistance in bacteria is becoming an increasingly serious problem for the global healthcare system, leading to a reduction in the effectiveness of previously curable infections and even making some infections incurable. The World Health Organization (WHO) has listed it as one of the biggest threats to public healthcare in the 21st century. To address this issue, significant research efforts are being invested into discovery of new antimicrobial drugs. Peptidoglycan is an important component of the bacterial cell wall, and its biosynthesis occurs in three regions: the cytoplasm, the cell membrane, and outside the cell. The MurA enzyme is present in the cytoplasm and initiates the pathway by catalyzing the first reaction. The majority of currently available antibiotics inhibit the peptidoglycan synthesis pathway, with their site of action in the later stages outside the cell. However, a few antibiotics act inside the cell, such as fosfomycin, which inhibits the activity of the MurA enzyme. This master’s thesis focused on synthesizing two glucosamine derivatives based on molecular modeling and mimicking of the natural substrate. N-acetylglucosamine was utilized to correctly align functional groups and develop reversible inhibitors capable of penetrating the cell wall and membrane, reaching the bacteria’s interior where the MurA enzyme is located. The primary purpose of the master’s thesis was to develop a synthetic route for preparing compounds that have acidic fragments attached to the hydroxyl group located at position 6. Firstly, all functional groups of N-acetylglucosamine, except for the hydroxyl group at position 6, were protected. Compound 11 was synthesized by directly introducing acetic acid onto the hydroxyl group using chloroacetic acid. For the synthesis of compound 9, the hydroxyl group was first activated by forming a tosylate, which was then utilized to form a derivative with dimethyl malonate. This derivative was finally hydrolyzed to yield carboxylic acid form. Enzyme testing confirmed that both compounds inhibited enzyme activity, with compound 9 having an IC50 value of 84 μM and compound 11 having an IC50 value of 14 µM.

• Keywords: antibiotic resistance, peptidoglycan, inhibitor, N-acetylglucosamine, MurA enzyme