Augustus, Brian W. J. (1980) Studies on sialidases. PhD thesis, University of Warwick.

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Abstract

The subject, "Studies On Sialidases", is introduced In Chapter 1 by a review of the history, biological functions and properties of sialidases.

The development of a rapid and sensitive (detection limit 10-⁶ units) assay system for sialidase, using tritiated fetuin labelled at C-8 of its N-acetylneuraminic acid (NANA) units, is discussed in Chapter 2. Included In this chapter, is the design and application of a radioimmunoassay for sialidases, in which antibodies specific for (NANA) have been obtained. These antibodies were raised in sheep challenged with BSA-colominic acid (the colominic acid was hydrolysed to a chain length of approximately 3 NANA units) conjugate. Further, an attempt has been made to determine the mole to mole ratio of NANA units per sialic acid-containing macromolecule.

The extensive purification of the three sialidases is discussed in Chapter 3, using a double "affinity" column system and hydroxylapatite with which it was possible to separate the two forms of sialidases found in S. griseus and V. cholerae.

The main body of the thesis (Chapter 4), reports on the work undertaken to resolve the controversy pertaining to which amino acids are, or are not, involved in the active centre of sialidases obtained from pathogenic (C. perfringens and V. cholerae) and non-pathogenic (S. griseus) sources. It was found, by chemical modification, that cysteine and lysine amino acid residues do not participate in the catalytic process, whereas on the other hand, arginine, tryptophan and carboxylic amino acid residues have been found to participate in or near the active centre, either by binding or by actual catalysis, during the catalytic process. Further, the involvement of arginine has been confirmed by differential labelling, isolation of the radiolabelled "active site peptide" and amino acid analysis, using phenyl(³H]glyoxal. The synthesis of the latter, via an inexpensive method, is elaborated upon as well. Preliminary chemical modification studies directed at the carbohydrate residues, covalently attached to these enzymes, reveal the possibility of these moieties participating in the maintenance of the three-dimensional structure of sialidase.

With the aforegoing information, it is concluded that there is no difference between the active centres of sialidases obtained from pathogenic and non-pathogenic bacterial sources. Further, a catalytic mechanism involving the essential amino acid residues in or near the active site of the sialidases is speculated upon.

Item Type:	Thesis or Dissertation (PhD)
Subjects:	Q Science > QD Chemistry
Library of Congress Subject Headings (LCSH):	Neuraminidase, Radioimmunoassay, Amino acids
Official Date:	August 1980
Dates:	Date Event August 1980 Submitted
Institution:	University of Warwick
Theses Department:	Department of Chemistry
Thesis Type:	PhD
Publication Status:	Unpublished
Supervisor(s)/Advisor:	Hutchinson, D. W. (David Wesley)
Sponsors:	Oppenheimer Memorial Trust
Format of File:	pdf
Extent:	v, 174 leaves : illustrations, charts
Language:	eng

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