Purification and Characterization of a Novel Intracellular Sucrase Enzyme ofLeishmania donovaniPromastigotes

Abstract

The promastigote stage of Leishmania resides in the sand fly gut, enriched with sugar molecules. Recently we reported that Leishmania donovani possesses a sucrose uptake system and a stable pool of intracellular sucrose metabolizing enzyme. In the present study, we purified the intracellular sucrase nearly to its homogeneity and compared it with the purified extracellular sucrase. The estimated size of intracellular sucrase is ~112 kDa by gel filtration chromatography, native PAGE, and substrate staining. However, in SDS-PAGE, the protein is resolved at ~56 kDa, indicating the possibility of a homodimer in its native state. The kinetics of purified intracellular sucrase shows its higher substrate affinity with a K m of 1.61 mM than the extracellular form having a K m of 4.4 mM. The highly specific activity of intracellular sucrase towards sucrose is optimal at pH 6.0 and at 30°C. In this report the purification and characterization of intracellular sucrase provide evidence that sucrase enzyme exists at least in two different forms in Leishmania donovani promastigotes. This intracellular sucrase may support further intracellular utilization of transported sucrose.

<p>Beneficial effects of Japanese sake yeast supplement on biochemical, antioxidant, and anti-inflammatory factors in streptozotocin-induced diabetic rats</p>

“…Treatment with sake yeast supplement improved GSH and TAS levels which may be due to decreased production of free radicals and increased level of antioxidants. Previous studies have reported that STZ causes diabetes by elevating sensitivity to lipid peroxidation and oxidative stress, thus pointing to mechanisms probably involved in the pathogenesis of the disease 9,21. MDA has been reported as one of the products of lipid peroxidation that is rapidly mixed with biomolecules and therefore disturbs the glucose metabolism 22.…”

“…Hypertriglyceridemia and hypercholesterolemia are known as common signs in diabetes 28. Studies have also reported that STZ increases the sensitivity to lipid peroxidation 9,29. Decreased hepatic insulin sensitivity could be attributed to increased hepatic gluconeogenesis, postprandial hyperinsulinemia, and increased production of triglycerides in the liver cells.…”

“…C-reactive protein (CRP) has also been reported to have a role in the initiation and aggravation of the classical pathways and promotion of atherosclerosis in diabetic patients 8. Imbalance between oxidant and antioxidant factors may result in oxidative stress which may increase the production of free radicals and reduces antioxidant factors 9. It is known that antioxidants are capable of removing free radicals and ROS by preventing lipid peroxidation and reducing the side effects that are caused by ROS 10.…”

Purification and biochemical characterization of β-d-fructofuranosidase fromBacillussubtilisLYN12

“…In the present study, significant inhibition by PMSF indicates the presence of the amino acid serine in the catalytic site. The inhibition by the thiol modifying reagent NEM depicts the presence of sulphydryl groups (‐SH) in the active site of β‐ d ‐fructofuranosidase (Singh & Mandal, ). The inhibition by TLCK indicates the possibility of histidine moieties, as well as the inhibition by NBS may indicate the presence of histidine/tryptophan/methionine amino groups in the catalytic triad.…”