Structure of peptide from active site region of Escherichia coli L-asparaginase.
The L-asparagine analogue 5-diazo-4-oxo-L-[5-14C]norvaline binds irreversibly to the active site of Escherichia coli L-asparaginase. Conditions for optimal labeling in buffers containing 50% dimethylsulfoxide have been developed and kinetic parameters of the inactivation have been determined. After reduction, alkylation and subsequent degradation of the modified enzyme with alpha-chymotrypsin, the principal radioactive decapeptide of sequence Val-Gly-Ala-Met-Arg-Pro-Ser-Thr-Ser-Met was isolated. A second radioactive hexapeptide Arg-Pro-Ser-Thr-Ser-Met resulting from chymotryptic digestion of the decapeptide was also isolated. Evidence is presented for the attachment of the 5-diazo-4-oxo-L-norvaline residue to serine-9 in the decapeptide via an acid-labile linkage.
D000595:Amino Acid Sequence; D001215:Asparaginase / Q000378:metabolism; D001216:Asparagine / Q000031:analogs & derivatives / Q000378:metabolism; D001665:Binding Sites; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D046911:Macromolecular Substances; D010446:Peptide Fragments / Q000032:analysis; D011485:Protein Binding
D046911:Macromolecular Substances; D010446:Peptide Fragments; D001216:Asparagine; D001215:Asparaginase; D004121:Dimethyl Sulfoxide