RESUMEN
We report the usefulness of yam tyrosinase as a catalyst in the synthesis of cross-linked protein networks for biopolymers. The enzyme was purified using aqueous two-phase partitioning (ATPs) and peptide mapping on SDS-PAGE was carried out to ascertain degree of similarities of tyrosinase from the yam species. The mapping revealed distinct peptide bands of 3, 4, 4 and 2 for tyrosinase from D. praehensilis, D. alata, D. rotundata and C. esculenta respectively purified using conventional method. In contrast, continuous broad band was noticed for the ATPS-purified enzymes due to bound polyethylene glycol (PEG). Tyrosinase from D. praehensilis with overall better properties was used in the synthesis of cross-linked protein networks. The enzyme catalyzed conversion of soluble proteins from whey, moringa leaves, pumpkin leaves and cow blood into fibrous (cross-linked) protein networks for improved properties and functionalities. The purified tyrosinase from D. praehensilis was also covalently bonded to bovine serum albumin (BSA) forming tyrosinase-BSA adduct with molecular weight of 118 ± 2.0 kDa, revealing its potential as a reporter enzyme by reporting BSA. The overall result further reinforces yam tyrosinase as an enzyme of interest in various biotechnological applications.
RESUMEN
A shortened method of purification and immobilization of tyrosinase from different species of yam (Dioscorea spp) on insoluble supports is described. The enzyme was purified by aqueous two-phase partitioning (ATPS) followed by gel filtration chromatography. The purified enzyme was immobilized on Ca-alginate, polyacrylamide gel or as cross-linked enzyme aggregate (CLEA) to obtain a yield of between 51-64%, 33-46% and 52-65% respectively for all the yam species. The optimum pH obtained for tyrosinase immobilized on polyacrylamide gel and CLEA was equivalent to that of free enzyme (pH 6.5). In contrast, Ca-alginate entrapped tyrosinase exhibited a shift of optimum pH to 7.0. Entrapped Tyrosinase in polyacrylamide gel and Ca-alginate also retained the same optimum temperature as the free enzyme (50 °C). While the optimum temperature of CLEA shifted to 60 °C. When subjected to four repeated use cycles, tyrosinase entrapped in polyacrylamide gel, Ca-alginate and CLEA still retained close to 40, 35 and 45 % of their initial activities respectively after the fourth cycle. The overall result further suggests yam tyrosinase as a promising enzyme for biocatalysis and biotechnological applications.
RESUMEN
BACKGROUND: The study determined the fractions of proteins in the urine and plasma of rats at different ages, measured the plasma and urine concentrations of markers of renal function, with a view to determining the influence of proteinuria on renal function. METHODS: Eighty Wistar rats were used for this study. Groups 1 and 2 each consisted of eight 1-month-old male and female rats; 3 and 4 had eight 3-month-old male and female rats; 5 and 6 had eight 6-month-old male and female rats; 7 and 8 had eight 9-month old male and female rats; and 9 and 10 had eight 12-month-old male and female rats. RESULTS: A fraction of the molecular weight of protein in the urine of rats aged 1, 9 and 12 months was higher than that of 3 and 6 months. The total protein concentration in the urine of male and female rats aged 9 and 12 months was significantly higher than that of rats aged 1 and 3 months. The urine creatinine concentrations of male and female rats aged 9 months were significantly higher when compared with that of 1, 3, 6 and 12 months. CONCLUSION: Our results suggest that the 3-month-old rats seem less affected by proteinuria, because they had the least urine protein, and consistent and reduced plasma and urine concentrations of markers of renal function. The results of this study may provide a foundation for future mechanistic inquiries as to why this age group was the least affected by proteinuria.