Kinetic analysis of papaya proteinase omega.
Papaya proteinase omega (pp omega) has been purified from dried latex both by immunoaffinity and traditional methods. Kinetic analysis revealed that (1), the pp omega-catalysed hydrolysis of N-benzoyl-L-arginine p-nitroanilide (BApNA) has a lower specificity (kcat/Km) than the same reaction catalysed by papain; (2), the pp omega-catalysed hydrolysis of a tripeptide substrate having phenylalanine at the second position (S2-site) showed a more similar specificity to that catalysed by papain; (3), the significant difference between the two enzymes is that steady state kinetics with both L-BApNA and a tripeptide enables the identification in pp omega of other ionizations affecting binding. The active sites of papain and pp omega can therefore be distinguished by pH-dependence of kcat/Km.
Protein Engineering Department, AFRC Institute of Food Research, Reading Laboratory, Reading, UK.
I G
IG
Sumner
A
A
Vaughan
R
R
Eisenthal
R W
RW
Pickersgill
A J
AJ
Owen
P W
PW
Goodenough
D000595:Amino Acid Sequence; D000906:Antibodies / Q000737:chemistry / Q000302:isolation & purification; D001586:Benzoylarginine Nitroanilide / Q000378:metabolism; D001665:Binding Sites; D003546:Cysteine Endopeptidases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D007840:Latex / Q000737:chemistry; D008969:Molecular Sequence Data; D010206:Papain / Q000737:chemistry; D010940:Plant Proteins
D000906:Antibodies; D007840:Latex; D010940:Plant Proteins; D001586:Benzoylarginine Nitroanilide; D003546:Cysteine Endopeptidases; C018745:actinidain; D010206:Papain; C039974:caricain