Cyclodextrins as templates for the presentation of protease inhibitors.
Mono(6-succinylamido-6-deoxy)-beta-cyclodextrin was synthesized by classical carbohydrate chemistry and used as a template mono-functionalized with the linear, fully flexible 4C-spacer carboxylate for covalent linkage of the calpain inhibitor leucyl-leucyl-norleucinal. Spectroscopic analyses of the conjugate do not support a self-inclusion of part of the hydrophobic peptide tail, but confirm its intra- or intermolecular interaction with the template moiety that leads to full water solubility. The inhibitory potency of the beta-cyclodextrin/peptide aldehyde construct was compared with that of the parent Ac-Leu-Leu-Nle-H against cathepsin B and calpain. Despite the large size of the template the inhibition of cathepsin B was only slightly reduced in full agreement with the X-ray structure of this enzyme which shows full accessibility of the S-subsites. For this enzyme the 4C-spacer is apparently sufficient to guarantee optimal interaction of the peptide tail with the binding cleft. Conversely, for mu-calpain a significantly decreased inhibitory potency was obtained with the conjugate suggesting steric interference of the template in the binding process. These results show that the beneficial properties of the cyclodextrin template can be retained in conjugates with bioactive peptides if attention is paid to optimize in each case the size and nature of the spacer for optimal recognition of the grafted biomolecule.
Schaschke
N
N
Max-Planck-Institut für Biochemie, AG Bioorganische Chemie, Martinsried, Germany.
Musiol
H J
HJ
Assfalg-Machleidt
I
I
Machleidt
W
W
Rudolph-Böhner
S
S
Moroder
L
L
D002154:Calpain / Q000037:antagonists & inhibitors*; D002240:Carbohydrate Sequence; D002942:Circular Dichroism; D003505:Cyclodextrins / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D007700:Kinetics; D007976:Leupeptins / Q000494:pharmacology*; D008969:Molecular Sequence Data; D047392:beta-Cyclodextrins
D003505:Cyclodextrins; D015853:Cysteine Proteinase Inhibitors; D007976:Leupeptins; D047392:beta-Cyclodextrins; C063261:acetylleucyl-leucyl-norleucinal; D002154:Calpain; C031215:betadex