Comparative Analysis of the Interaction of Silver Nanoparticles with Hexokinase from Trypanosoma brucei and Humans

Regardless of the efforts to ease cases of human African trypanosomiasis (HAT), an increased number of cases get reported annually. This is because of drug resistant Trypanosoma brucei (Tb), the causative agent of the illness. This has renewed the need for creative methods to find new anti-trypanosomal drugs. The blood stream form (BSF) of the parasite depends exclusively on the glycolytic pathway for energy production while it is in the human host. Interruptions in this pathway efficiently kills the parasite. Trypanosoma brucei hexokinase (TbHK) is the first enzyme in glycolysis, and any effectors or inhibitors of TbHK would have potential as anti-trypanosomal agents.

TbHK and human glucokinase (hGCK) were over-expressed with a 6 histidine-tag in E. coli BL21(DE3) cells having the pRARE2 plasmid.

TbHK had thermal and pH stability between 30°C and 55°C and 7.5 and 8.5, respectively, while hGCK exhibited thermal and pH stability between 30°C and 40°C and 7.0 and 8.0, respectively. Kinetically, TbHK had a K_m of 39.3 µM, V_max of 0.066 µmol.min^-1.mL^-1, k_cat of 2.05 min^-1 and k_cat/K_m of 0.0526 min^-1.µmol^-1. hGCK exhibited a K_m of 4.5 µM, V_max of 0.032 µnmol.min^-1.mL^-1, k_cat of 11.25 min^-1, and k_cat/K_m of 2.5 min^-1.µmol^-1. Interaction kinetic studies of silver nanoparticles (AgNPs) (0.1 µM) of average size of 6 nm with TbHK and hGCK were conducted. AgNPs selectively inhibited TbHK over hGCK. TbHK showed a non-competitive inhibition with a 50% and 28% decrease in V_max, and k_cat/k_m, respectively. HsGCK showed a 33% increase in affinity, 9% decrease in V_max, and a 50% increase in enzyme efficiency.

The observed pattern of hGCK and AgNPs falls under the uncompetitive inhibition. The observed highly selective inhibitory effects of AgNPs between TbHK and hGCK may be used in development of new anti-trypanosomal drugs.