ABSTRACT
Phosphoglucose isomerase (EC 5.3.1.9) catalyzes the interconversion of D-glucopyranose-6-phosphate and D-fructofuranose-6-phosphate by promoting an intrahydrogen transfer between C1 and C2. A conserved histidine exists throughout all phosphoglucose isomerases and was hypothesized to be the base catalyzing the isomerization reaction. In the present study, this conserved histidine, His311, of the enzyme from Bacillus stearothermophilus was subjected to mutational analysis, and the mutational effect on the inactivation kinetics by N-bromoacetylethanolamine phosphate was investigated. The substitution of His311 with alanine, asparagine, or glutamine resulted in the decrease of activity, in k(cat)/K(M), by a factor of 10(3), indicating the importance of this residue. N-bromoacetylethanolamine phosphate inactivated irreversibly the activity of wild-type phosphoglucose isomerase; however, His311 --> Ala became resistant to this inhibitor, indicating that His311 is located in the active site and is responsible for the inactivation of the enzyme by this active site-directed inhibitor. The pKa of His311 was estimated to be 6.31 according to the pH dependence of the inactivation. The proximity of this value with the pKa value of 6.35, determined from the pH dependence of k(cat)/K(M), supports a role of His311 as a general base in the catalysis.
Subject(s)
Enzyme Inhibitors/pharmacology , Ethanolamines/pharmacology , Geobacillus stearothermophilus/enzymology , Glucose-6-Phosphate Isomerase/chemistry , Glucose-6-Phosphate Isomerase/metabolism , Histidine , Amino Acid Sequence , Amino Acid Substitution , Animals , Arabidopsis/enzymology , Binding Sites , Conserved Sequence , Enzyme Inhibitors/chemistry , Escherichia coli/enzymology , Ethanolamines/chemistry , Glucose-6-Phosphate Isomerase/antagonists & inhibitors , Humans , Kinetics , Mice , Molecular Sequence Data , Mutagenesis, Site-Directed , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Saccharomyces cerevisiae/enzymology , Zymomonas/enzymologyABSTRACT
A survey of Schistosoma mansoni infection and snail hosts was carried out in 1992 in six accessible schools and 11 water bodies respectively. Five to ten per cent of the students were randomly selected for stool examination by the Ritichie's method. In Adwa town, however, stools of 100 students out of the 199 selected were re-examined by Kato's method to assess intensity of infection. Results of stools examined by Ritichie's method showed that S. mansoni positive patients were present in four out of six (66.7%) schools, the prevalence for school ranging from 1% in Maychew to 61.8% in Adwa. In all the localities, there was significant male preponderance in the prevalence of infection (p < 0.05). Among those whose stools were examined by the Kato's method in Adwa town, both prevalence and geometric mean egg count per gram of faeces (EPG) were highest in the 10 to 14 years age group reaching 68% and 597 followed by 64% and 591 respectively in the 15 to 19 years of age. Of the S. mansoni positive students in Adwa, 86% excreted 200 or more EPG, 84% of those in the 10 to 14 years of age excreted as high as 1,600 EPG. Sexwise, no significant difference was observed in EPG (p > 0.05). Biomphalaria pfeifferi, the principal snail host of S. mansoni in Ethiopia, were collected from five out of the 11 waterbodies and were found shedding human schistosome cercariae in two of them, their infection rate reaching 0.7%. The ecological characteristics observed were found suitable for snail propagation.(ABSTRACT TRUNCATED AT 250 WORDS)
