The histidine specific reagent 2-phenyl-l,4-dibromoacetoin (PDA) has been applied to αSI-casein B. Reaction of αSI-casein with PDA for 26 hours resulted in the loss of about two residues each of histidine and methionine per αSI-casein monomer (molecular weight = 27,000). The modified protein was 90% soluble in 8 mM calcium chloride and precipitated quantitatively at 13 mM calcium chloride. The control was precipitated quantitatively at 8 mM. The calcium binding capacity of the modified αSI-casein was reduced to about 4.5 calcium ions per PDA αsi-casein monomer from 12.4 calcium ions per αSl-casein monomer. Reaction of αSI-casein with PDA resulted in the production of aggregated material which remained at the origin on polyacrylamide gel electrophoresis in the presence of urea and 2-mercaptoethanol and which was eluted at the void volume of a Sephadex G-200 column.
While following the time course of the reaction of PDA with αSI-casein, it was found by N-bromosuccinimide (NBS) spectrophotometric titration (6 M urea-acetate-formate buffer, pH 4.0) that two residues of tryptophan per monomer αSI-casein could be detected at zero reaction time but 3.1 residues could be detected after 30 hours. Comparison of the results obtained with αSI-casein, PDA αSl-casein, β-lactoglobulin and α-chymotrypsinogen using both the NBS titration procedure and the p-dimethylaminobenzaldehyde method of Spies and Chambers (46) suggest the presence of a "buried" tryptophan residue in αSI-casein. PDA has a negligible effect on the NBS titration procedure so the residue "exposed" cannot be an artifact generated by PDA bound to the protein. Increasing the urea concentration to 10 M did not expose the third tryptophan residue to NBS. / Land and Food Systems, Faculty of / Graduate
Identifer | oai:union.ndltd.org:UBC/oai:circle.library.ubc.ca:2429/34462 |
Date | January 1970 |
Creators | Beveridge, Herbert James Thomas |
Publisher | University of British Columbia |
Source Sets | University of British Columbia |
Language | English |
Detected Language | English |
Type | Text, Thesis/Dissertation |
Rights | For non-commercial purposes only, such as research, private study and education. Additional conditions apply, see Terms of Use https://open.library.ubc.ca/terms_of_use. |
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