Date of Award

1982

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Abstract

This report contributes to the knowledge of antiencephalitogenic spinal cord protein (SCP) with respect to three major areas of study: (1) the purification and characterization of human SCP; (2) the nature of the association of SCP with nervous tissue components and (3) purification of cyanogen bromide-derived peptides of bovine SCP.;Human SCP and a protein immunochemically identical to SCP (SCP-PN) were purified from spinal cords and peripheral nerves with 0.15 M sodium chloride, carboxy-methyl cellulose chromatography and gel filtration on Sephadex G-50 superfine. SCP and SCP-PN had estimated molecular weights of 13,700 and 14,700 daltons, respectively and had similar amino acid compositions. The isoelectric point of SCP-PN was estimated to be 9.9. Immunodiffusion analyses with anti-human SCP sera or anti-bovine SCP sera revealed that human SCP and SCP-PN are each composed of two different antigenic forms. Each antigenic form contains a distinct immunogenic domain that is identical to one of the immunogenic sites on bovine SCP.;Bovine SCP-PN is identical to the P(,2) protein found in purified peripheral nerve myeline. The bovine SCP-PN content of 0.3 M NaCl extracts of whole tissue was 1.3 mg per g of tissue. Approximately 0.33 mg of SCP-PN was found in the soluble fraction of 0.8 M sucrose homogenates of bovine peripheral nerves. Densitometry data indicated that SCP-PN decreased from 19% of the total myelin protein to less than 1% when purified myelin was extracted with 0.3 M sodium chloride or 0.05 M hydrochloric acid. The basic proteins SCP-PN and lysozyme bound to myelin and sodium chloride-extracted myeline when they were added to a suspension of myelin in 0.8 M sucrose. Pepsin, an acidic protein, did not bind to myelin. The results suggest that in 0.8 M sucrose, positively charged SCP-PN can bind to negatively charged myelin. Myelin-associated SCP-PN behaves like a peripheral membrane protein.;This interpretation is consistent with earlier research in which bovine SCP-PN was localized by immunohistological techniques in axons of peripheral nerves but not in myelin sheaths surrounding the axons. The histological fixatives acetone and 95% ethanol/ether did not render SCP-PN in whole tissue insoluble in saline. This result indicates that if a tissue section is washed with saline after fixation with acetone or 95% ethanol/ether, then most of the SCP-PN in the tissue section could be solubilized. . . . (Author's abstract exceeds stipulated maximum length. Discontinued here with permission of school.) UMI

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