Tryptic cleavage as a probe of conformational differences between active and inactive forms of ornithine aminotransferase.
Treatment of ornithine aminotransferase with trypsin resulted in rapid and complete loss of enzyme activity in a process that coincided with a reduction in subunit Mr of about 3000. However, electrophoresis showed that a substantial proportion of the enzyme had not been digested. The component of the preparation of ornithine aminotransferase that was insusceptible to trypsin contained a naturally occurring but enzymically inactive form of the enzyme, and when this had been removed, the remaining fully active enzyme was completely digested. Irreversible inactivation with a substrate analogue made all of the enzyme insusceptible to trypsin. The hydrolyzed enzyme still underwent a very slow half-reaction with ornithine. Sequence analysis of the truncated protein, prepared by blotting from electrophoretic gels, showed that hydrolysis had occurred at peptide bond Lys26-Tyr27.
Simmaco
M
M
Department of Biochemistry, University College, Cardiff, Wales, United Kingdom.
Carr
L
L
Bossa
F
F
Barra
D
D
Basford
J M
JM
John
R A
RA
D000595:Amino Acid Sequence; D000818:Animals; D004789:Enzyme Activation; D007700:Kinetics; D008969:Molecular Sequence Data; D009952:Ornithine / Q000378:metabolism; D009953:Ornithine-Oxo-Acid Transaminase / Q000037:antagonists & inhibitors / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis; D011487:Protein Conformation; D051381:Rats; D000637:Transaminases / Q000037:antagonists & inhibitors / Q000378:metabolism; D014357:Trypsin / Q000494:pharmacology*
D010446:Peptide Fragments; D009952:Ornithine; D000637:Transaminases; D009953:Ornithine-Oxo-Acid Transaminase; D014357:Trypsin