Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.773381
Title: Peptide hydrolases of the brush border membrane from human small intestine
Author: Sterchi, Erwin E.
Awarding Body: Keele University
Current Institution: Keele University
Date of Award: 1978
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Abstract:
A method has been developed for the purification of brush border membranes from human small intestine. The method did not involve the use of EDTA-buffers or disruption of brush borders with high concentrations of Tris. On average, a 24-fold increase in specific activity for ot-giucosidase (brush border marker) was obtained in the final preparation which contained only traces of enzyme markers from other cellular organelles. The homogenates of human small intestinal mucosa were shown to contain enzymes capable of hydrolysing di-, tri- and oligopeptides as well as 2-naphthylamides. Distribution studies indicated that all of the oligopeptidase activity towards peptides, four and more amino acids in length and activity towards leucine-2 -naphthylamide were located exclusively in the brush border. A large proportion of activity towards a-glutamic acid-2-naphthylamide (aminopeptidase A), y-glutamic acid-2 -naphthylamide (y-glutamyltransferase) and glycyl- proline-2 -naphthylamide (dipeptidyl peptidase IV) were also recovered in the brush border membrane fraction. Depending on the substrate used, 33-87% of tripeptidase activity was located in the brush border membrane. An estimated 58-87% of dipeptidase activity, on the other hand, was recovered in the soluble fraction. Solubilisation of brush border membrane proteins by sodium dodecyl sulphate, Triton X-100 and papain followed by polyacrylamide gel electrophoresis revealed seven different peptide hydrolases. These included the specific enzymes aminopeptidase A, dipeptidyl peptidase IV, y-glutamyltransferase and aminopeptidase M which were clearly separable on polyacrylamide gels after solubilisation with Triton X-100 or papain. Activity recovered in the aminopeptidase M peak in the above gel system could be resolved into two distinct peptidases in addition to aminopeptidase M, by SDS-gel electro­phoresis. One of these peptidases was most active towards aliphatic tripeptides (peptidase 1 ) while the other appeared to be specific for dipeptides. A further peptidase (aminopeptidase M') was resolved by isoelectric focusing.in polyacrylamide gels. The role of these brush border peptide hydrolases in the absorption of protein by the gut is discussed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.773381  DOI: Not available
Keywords: QH301 Biology
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