Mechanism of inhibition of papain by chicken egg white cystatin. Inhibition constants of N-terminally truncated forms and cyanogen bromide fragments of the inhibitor.
N-terminally truncated forms of chicken egg white cystatin and its cyanogen bromide fragments were isolated and assayed for inhibition of papain. Truncated forms beginning with Gly-9 and Ala-10 had a 5000-fold lower affinity for papain than the two isoelectric forms (pI = 6.5 and 5.6) of the full-length inhibitor (Ki = 6 pM and 7 pM) or a truncated form beginning with Leu-7 (Ki = 6 pM), indicating the outstanding importance of one or two residues preceding conserved Gly-9 for binding. A weak inhibition of papain (Ki = 900 nM) was exhibited by the intermediate cyanogen bromide fragment (residues 30-89) containing the chicken cystatin QLVSG variation of the QVVAG segment which is conserved in almost all members of the cystatin superfamily. The obtained affinity data provide independent evidence for the validity of the proposed docking model of a chicken cystatin-papain complex [(1988) EMBO J. 7, 2593-2599].
Machleidt
W
W
Institut für Physiologische Chemie, Universität München, FRG.
Thiele
U
U
Laber
B
B
Assfalg-Machleidt
I
I
Esterl
A
A
Wiegand
G
G
Kos
J
J
Turk
V
V
Bode
W
W
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002645:Chickens; D002845:Chromatography / Q000379:methods; D003488:Cyanogen Bromide; D015891:Cystatins; D007525:Isoelectric Focusing; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010206:Papain / Q000037:antagonists & inhibitors*; D010446:Peptide Fragments / Q000494:pharmacology; D011506:Proteins / Q000494:pharmacology*
D015891:Cystatins; D010446:Peptide Fragments; D011506:Proteins; C100158:cystatin, egg-white; D010206:Papain; D003488:Cyanogen Bromide