Peptide-Based Drug Design

60 Winkler and Campbell
amino acids defined. After the first synthesis and screening round there are two
possibilities: Either one can combine different sequence patterns to a complete
sequence (56,57) or choose the sequence pattern with the strongest activity. The
next combinatorial library is assembled with the sequence pattern conserved
and the deconvolution of the other one or two mixture positions. This repeating
of deconvolution, synthesis, and screening cycles proceeds until one obtains a
complete sequence in which all amino acid positions are defined (43,48,54).
A special type of combinatorial library is the positional scanning library (43,
49,58). This array contains a set of combinatorial libraries with one or more (two
positions = dual positional scan (48,59))deconvoluted positions. In each of the
synthesized libraries the deconvoluted positions are different. After screening
one can combine the positions of the spot with the peptides of highest activity
of each library to obtain the complete sequence of a peptide with most likely the
greatest activity.
In order to reduce the number of possible combinations, combine several
amino acids of similar properties in clusters (e.g., hydrophobicity—isoleucine,
leucine, valine; or steric similarity—serine, cysteine, aminobutyric acid). These
clustered amino acid peptide libraries provide easier and faster screening of large
peptide libaries (60). It is of course necessary to resolve these clusters at the end
of the screening process.
3.2.4. Others
3.2.4.1. RANDOM PEPTIDE LIBRARY
Another possibility to screen for active peptides without prior knowledge
of a starting sequence is use of a random peptide library. This array contains
randomized peptide sequences (61). Compared with the combinatorial peptide
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Fig. 5. (Continued) �-amino group and the carboxy side chain function of the Cterminal
glutamic acid. In the first library only two positions of the peptide sequence
were defined (B1 in rows and B2 in columns; cysteine was omitted). At all other
positions, mixtures of all common l-amino acids were coupled (X). From the resulting
pattern, the strongest spot was chosen for defining the sequence motif for the next
screening cycle (B1 = FandB2 = N; see arrow). At the second screening cycle two of
the positions with mixtures were deconvoluted, now (B1, B2). The sequence motif FN
was synthesized in all peptide sequences. After probing, one of the strongest signals was
chosen for the extended sequence motif (B1 = HandB2 = D, resulting in HFND; see
arrow). With this sequence motif the third coupling cycle was performed to determine
the remaining undefined positions. In this screening cycle each spot contains one single
peptide with a defined sequence.