Protein Protocols Protein Protocols

1070 Ehrlich, Berthold, and Bailon
11. The 32 codons encoding the possible 20 amino acids are (1) TTT = F (phenylalanine),
(2) TTG = L (leucine), (3) TCT = S (serine), (4) TCG = S, (5) TAT = Y (tyrosine), (6)
TAG = Q (glutamine), (7) TGT = C (cysteine), (8) TGG = W (tryptophan), (9) CTT = L
(leucine), (10) CTG = L, (11) CCT = P (proline), (12) CCG = P, (13) CAT = H (histidine),
(14) CAG = Q (glutamine), (15) CGT = R (arginine), (16) CGG = R, (17) ATT = I (isoleucine),
(18) ATG = M (methionine), (19) ACT = T (threonine), (20) ACG = T, (21)
AAT = N (asparagine), (22) AAG = K (lysine), (23) AGT = S, (24) AGG = R (arginine),
(25) GTT = V (valine), (26) GTG = V, (27) GCT = A (alanine), (28) GCG = A, (29) GAT = D
(aspartic acid), (30) GAG = E (glutamic acid), (31) GGT = G (glycine), and (32) GGG = G.
12. Biopanning is an ideal way of selecting for phage displayed binding peptides. However, it
is important that substrate specificity for selected phage is determined, as not all phage are
always binding phage. Some of these nonspecific binding phage might be plastic binders
(37) or they might be favored by natural selection (see Note 13) during phage amplication.
Although phage selectivity can be built directly into the target/phage interaction (38), the
most common and fastest way of determining substrate specificity is by enzyme-linked
immunosorbent assay (ELISA) using an antiphage antibody (27,39).
13. The “quickscreen” has been employed to avoid this problem, as no phage amplification
occurs between rounds of biopanning, that is, recovered phage from the first round is used
for the second round upon elution, and so forth (5). In this method, stepwise pH elution
can be used to select for high-affinity binding phage (5).
Acknowledgment
We appreciate the assistance of G. Katarina Luhr (Karolinska Institute, Stockholm,
Sweden) in rendering Fig. 1.
References
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antigens on the virion surface. Science 228, 1315–1317.
2. Braisted, A. C. and Wells, J. A. (1996) Minimizing a binding domain from protein A.
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Binding proteins selected from combinatorial libraries of an alpha-helical bacterial receptor
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and Applications of Biomolecular Recognition. Abstract L30, Kalmar, Sweden.
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region of a humanized IgG1 monoclonal antibody using phage display. J. Mol. Recogn. 11,
121–125.
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