Enzyme activity and AGE formation in a model of AST glycoxidation by D-fructose in vitro

Scientific paper - Preliminary note / Short communication

• Title (croatian): Katalitička aktivnost i stvaranje AGE-produkata u modelu glikoksidacije AST D-fruktozom in vitro
• Author: Iva Bousova
• Author: Danka Vukasovic
• Author: Dubravka Juretic
• Author: Vladimir Palicka
• Author: Jaroslav Drsata
• Author's institution: University of Zagreb; Faculty of Pharmacy and Biochemistry; (Department of medical biochemistry and haematology)
• Scientific / art field, discipline and subdiscipline: BIOMEDICINE AND HEALTHCARE; Pharmacy; Medical Biochemistry

Abstract (english)

Non-enzymatic glycation as the reaction chain between reducing sugars and free amino groups of proteins has been shown to correlate with physiological ageing and severity of diabetes. The process involves oxidative steps (glycoxidation). In this paper, the effect of D-fructose as a reactive sugar on aspartate aminotransferase (AST) as a model protein was monitored by measurements of the enzyme activity and formation of fluorescent advanced glycation end products (AGEs). Change in the AST activity was considered as a measure of the overall protein damage caused by glycation, and total AGEs and pentosidine represent, at least partly, the formation of glycoxidation products. Catalytic activity of AST in an incubation mixture containing D-fructose (50 mmol L-1), decreased compared to control values to 42% (p < 0.05) and to 11% (p < 0.05) on 5th and on 21st day of incubation, respectively. In the presence of fructose, total fluorescent AGEs concentrations were significant higher since 5th day of incubation (110 %, p < 0.05) and the fluorescent pentosidine concentrations from 15th day of incubation (117 %, p < 0.05) compared to control values, respectively. Catalytic activity of AST clearly and quantitatively demonstrated functional changes in the enzyme molecule caused by structural modifications initiated by fructose, while the evaluation of AGE formation and especially that of pentosidine by fluorescence measurement was less reliable.

Abstract (croatian)

Neenzimska glikacija je proces kojim se reducirajući šećeri kovalentnim vezama povezuju s amino skupinama proteina, bez posredovanja enzima. Taj proces dijelom je i oksidativan pa se često naziva glikooksidacija. Konačni produkti glikacije (AGE) nastaju polimerizacijom mnogih intermedijarnih produkata. U ovom radu, praćen je u in vitro uvjetima, mjerenjem katalitičke aktivnosti enzima i fluorescencije nastalih ukupnih AGE produkata i pentozidina, utjecaj D-fruktoze kao reaktivnog šećera na aspartat aminotransferazu (EC 2.6.1.1.) kao modelni protein. Promjena katalitičke aktivnosti enzima pokazatelj je promjenjene funkcije proteina glikacijom dok je intenzitet fluorescencije ukupnih AGE produkata i pentozidina samo djelomičan pokazatelj glikoksidacijskih promjena. Katalitička aktivnost AST izmjerena IFCC preporučenom metodom u inkubacijskoj smjesi koja je sadržavala 50 mmol L-1 fruktoze, smanjena je na 42% (p<0.05) petog dana, te na 11% (p<0.05) dvadeset i prvog dana. Statistički su značajne razlike u katalitičkoj aktivnosti enzima inkubiranog sa i bez fruktoze već nakon petog dana.Stvaranje ukupnih AGE produkata i pentozidina statistički je značajno nakon pet, odnosno 15 dana inkubacije enzima s fruktozom. Katalitička aktivnost enzima jasno ukazuje na funkcionalne promjene uzrokovane glikiranjem, dok je evaluacija stvaranja ukupnih AGE produkata, a pogotovo pentozidina, mjerenjem fluorescencije manje pouzdan pokazatelj promjena.

• Keywords (english): non-enzymatic glycation; glycoxidation; aspartate aminotransferase; AGE; pentosidine
• Keywords (croatian): neenzimska glikacija; glikooksidacija; aspartat aminotransferaza; krajnji produkti glikacije; pentozidin
• Language: english
• Language: croatian
• Publication type: Scientific paper - Preliminary note / Short communication
• Publication status: Published
• Peer review: Peer review - international
• Publication version: Published version
• Journal title: Acta Pharmaceutica
• Numbering: vol. 55, no. 1, pp. 107-114
• p-ISSN: 1330-0075
• e-ISSN: 1846-9558
• URN:NBN: urn:nbn:hr:163:074622
• Publication: 2005
• Document URL: https://hrcak.srce.hr/16749
• Type of resource: Text
• Access conditions: Open access
• Created on: 2020-04-08 14:55:56