| Abstract: | Earlier work has shown that 5-diazo-4-oxo-L-norvaline (DONV) irreversibly inactivates the L-asparaginase from E. coli by formation of a covalent bond in the region of the active site. Model compounds have been prepared to study this acid-labile covalent bond tentatively assigned to a serine or possibly a threonine residue in a decapeptide isolated from 14C-DONV-inactivated enzyme. Appropriately blocked DONV was found to alkylate methanol, and the hydroxyl function of blocked serine or threonine in the presence of boron trifluoride. The labile beta-ketoethers thus formed were reduced to the more stable beta-hydroxyethers. Facile lactonization of these 5-substituted-4-hydroxy-L-norvalines was observed. The diastereoisomers of both the lactonized and open forms of 5-methoxy-4-hydroxy-L-norvaline and related 4-hydroxy-L-2-amino acids of similar length were distinguishable on the amino acid analyzer. The beta-hydroxyethers derived from serine and threonine were hydrolyzed with acid and yielded the expected cleavage products. When the beta-ketoether was reduced by sodium borohydride prior to deblocking, in addition to the beta-hydroxyether, N-blocked amino alcohols were also formed, yielding a complex mixture of products. |
