Screening a random pentapeptide library, composed of 14 D-amino acids, against the COOH-terminal sequence of fructose-1,6-bisphosphate aldolase from Trypanosoma brucei.
A random pentapeptide library composed of 14 D-amino acids, including two unusual amino acids, thus representing 537,824 different peptide sequences anchored on polystyrene beads was created with each bead bearing a single pentapeptide sequence. This library was used for affinity screening against the fructose-1, 6-bisphosphate aldolase of Trypanosoma brucei labeled with biotin as well as versus the COOH-terminal labeled with fluorescein isothiocyanate. The thus selected peptide beads were identified and the appropriate sequences synthesized as peptide amides and evaluated for enzyme activity inhibition. Screening against the whole enzyme did not result in selection of an enzyme inhibitor. However, we demonstrate here that screening against a part of the enzyme involved in the catalytic activity may lead to the discovery of an enzyme inhibitor as well as an enzyme activator. Two low affinity inhibitors, RRVKF-NH2 and KThiKAR-NH2, with an IC50 of approximately 1 mM and approximately 0.2 mM, respectively, were identified. Two other pentapeptides with the sequence SWChaKK-NH2 and SKChaKM-NH2 are able to activate the enzyme fructose-1, 6-bisphosphate aldolase. Thus, successful screening of solid phase libraries can be accomplished using selected sequences of the target enzyme.
Samson
I
I
Laboratory of Medicinal Chemistry (F. F. W.), Rega Institute for Medical Research, Katholieke Universiteit Leuven, Minderbroedersstraat 10, B-3000 Leuven, Belgium.
Rozenski
J
J
Samyn
B
B
Van Aerschot
A
A
Van Beeumen
J
J
Herdewijn
P
P
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D001665:Binding Sites; D004305:Dose-Response Relationship, Drug; D005634:Fructose-Bisphosphate Aldolase / Q000037:antagonists & inhibitors*; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000494:pharmacology*; D019151:Peptide Library; D013237:Stereoisomerism; D014344:Trypanocidal Agents / Q000494:pharmacology; D014346:Trypanosoma brucei brucei / Q000201:enzymology*
D000596:Amino Acids; D009842:Oligopeptides; D019151:Peptide Library; D014344:Trypanocidal Agents; D005634:Fructose-Bisphosphate Aldolase