Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.26 (invertase)
 4,927 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A procedure was developed for the analytical isolation of brush border and basal lateral plasma membranes of intestinal epithelial cells. Brush border fragments were collected by low speed centrifugation, disrupted in hypertonic sorbitol, and subjected to density gradient centrifugation for separation of plasma membranes from nuclei and core material. Sucrase specific activity in the purified brush border plasma membranes was increased fortyfold with respect to the initial homogenate. Basal lateral membrane were harvested from the low speed supernatant and resolved from other subcellular components by equilibrium density gradient centrifugation. Recovery of Na, K-ATPase activity was 94%, and 61% of the recovered activity was present in a single symmetrical peak. The specific activity of Na, K-ATPase was increased twelvefold, and it was purified with respect to sucrase, succinic dehydrogenase, NADPH-cytochrome c reductase, nonspecific esterase, beta-glucuronidase, DNA, and RNA. The observed purification factors are comparable to results reported for other purification procedures, and the yield of Na, K-ATPase is greater by a factor of two than those reported for other procedures which produce no net increase in the Na, K-ATPase activity. Na, K-ATPase rich membranes are shown to originate from the basal lateral plasma membranes by the patterns of labeling that were produced when either isolated cells or everted gut sacs were incubated with the slowly permeating reagent 35S-p-(diazonium)-benzenesulfonic acid. In the former case subsequently purified Na, K-ATPase rich and sucrase rich membranes are labeled to the same extent, while in the latter there is a tenfold excess of label in the sucrase rich membranes. The plasma membrane fractions were in both cases more heavily labeled than intracellular protein. Alkaline phosphatase and calcium-stimulated ATPase were present at comparable levels on the two aspects of the epithelial cell plasma membrane, and 25% of the acid phosphatase activity was present on the basal lateral membrane, while it was absent from the brush border membrane. Less than 6% of the total Na, K-ATPase was present in brush border membranes.
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PMID:Analytical isolation of plasma membranes of intestinal epithelial cells: identification of Na, K-ATPase rich membranes and the distribution of enzyme activities. 13 16

1. In native invertase at pH 4.6, 23 out of a total of 34 tryptophyl residues are "exposed" to oxidation with N-bromosuccinimide, the other residues being apparently shielded from the oxidant within the molecule. 2. Oxidation of 5-6 tryptophyl residues/molecule with N-bromosuccinimide is proportional to the complete inactivation of the enzyme, and appears to be specific for indole chromophore only. The ligand binding and fluorescence measurements indicate that the oxidation of native enzyme, up to 50% inhibition, apparently does not change the conformation and topography of enzymes surface. 3. Invertase is inhibited by diazonium-1-H-tetrazole. Since tyrosine residues can be excluded by nitration studies as catalytically unimportant, it appears that a mocification of a single histidyl residue/molecule with diazonium-1-H-tetrazole is sufficient to abolish the enzymic activity. However, the absence of inhibition with diethyl pyrocarbonate indicates that the inhibition with diazonium-1-H-tetrazole may be mediated through steric hindrance or other indirect effects. 4. The absence of inhibition with 2,4-dinitrophenylhydrazine, trinitro benzenesulfonic acid and 5,5'-dithiobis-(2-nitrobenzoate) indicates that the carbonyl groups of the carbohydrate moiety, free amino and -SH groups are not essential for activity.
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PMID:External yeast beta-fructosidase. The role of tryptophyl residues in catalysis. 110 63