Inhibition Study Of GQDs And Trypanosoma Brucei Reductase

Trypanosoma is a protozoan worm that causes disease by parasitising the blood of a variety of animals.Of these,Trypanosoma brucei in Africa and Trypanosoma cruzi in South America are pathogenic to humans.Since the beginning of the 20 th century,human African trypanosomiasis has caused millions of deaths and poses a serious threat to human health.Nanomaterials are highly regarded in the scientific community due to their unique physicochemical properties and are now used in a variety of applications.Among them are graphene quantum dots(GQDs),which have been the focus of scientific attention since their discovery.It is a size-tunable and fluorescent carbon nanomaterial that is now commonly used in biomedical applications.Our previous studies have shown that GQDs can induce oxidative stress by affecting the energy metabolism of Trypanosoma brucei,leading to the continuous accumulation of reactive oxygen species in Trypanosoma brucei and eventually causing apoptosis of Trypanosoma brucei.This suggests that redox homeostasis plays an important role in the growth of Trypanosoma brucei.Trypanothione reductase of Trypanosoma brucei is specifically present in Trypanosoma brucei and this reductase plays a key role in controlling the redox homeostasis of the parasite,Do GQDs then interact with trypanothione reductase and have an effect on the life activities of the worm? If it can have an impact,what is the target of its impact? This thesis therefore provides a preliminary study of this.In this study,GQDs nanomaterials were first synthesized by chemical oxidation in the size range 2.7-4.4 nm,with a maximum emission peak at 440 nm.A recombinant expression plasmid containing a His tag to express trypanothione reductase was also constructed,and the corresponding recombinant protein was induced to be expressed and purified.The affinity of GQDs to trypanothione reductase was determined with the aid of a bio-layer interferometer,and it was found that GQDs had a strong affinity and a high binding amount to trypanothione reductase.Enzyme kinetic analysis was then carried out to measure the activity and Km of trypanothione reductase using trypanothione and NADPH as substrates.The inhibitory effect of GQDs on trypanothione reductase activity and the inhibition constants were determined using GQDs as inhibitors.9.024×102±0.003μmol·min-1mg-1 was measured and the Km value of the enzyme was 0.50 ± 0.02 μM.GQDs showed significant inhibition of trypanothione reductase activity at 10 μg.8)L-1 reconstituted trypanothione reductase had an IC50 value of19.921 μg.8)L-1 and an inhibition constant(Ki)of 0.0128 mg.8)L-1.This suggests that GQDs have a significant inhibitory effect on trypanothione reductase activity and that the inhibition is non-competitive.In summary,this study found that GQDs have a strong affinity for trypanothione reductase and inhibit the biological effects of Trypanosoma brucei by targeting this reductase to inhibit the viability of the worm.This study provides new research ideas for assessing the biological effects of GQDs and lays the foundation for the development of novel anti-trypanosome drugs.