Smith and Petrenko, Chemical Reviews, 1997, 97, 391-410

1
Smith and Petrenko, Chemical Reviews, 1997, 97,
391-410
2
Ligand bound to solid support
Assumption only tight binders remain bound to
solid support after washing steps
Urvoas, Minard and Soumillion Protein Engineering
by Phage Display in Protein Engineering Handbook,
2009, p563-603
3
The largest phage libraries have 1010
clones. Number of different random sequences
possible for 6-mers 6.4 x 107 15-mers 3 x
1019
Smith and Petrenko, Chemical Reviews, 1997, 97,
391-410
4

• Many different types of proteins can be displayed
  on phage. This allows for in vitro evolution.
• Optimize binding to target
• Optimize enzyme activity (?)

Smith and Petrenko, Chemical Reviews, 1997, 97,
391-410
5
Search sequence space for a more stably folded
protein.
Urvoas, Minard and Soumillion Protein Engineering
by Phage Display in Protein Engineering Handbook,
2009, p563-603
6
Activity-based screening of clones
Urvoas, Minard and Soumillion Protein Engineering
by Phage Display in Protein Engineering Handbook,
2009, p563-603
7
Searching sequence space for fitness
(fitness some desired property of the
mutagenized protein, which is used as a basis for
selection of clones)
Smith and Petrenko, Chemical Reviews, 1997, 97,
391-410
8
Crameri et al., Nature, 1998, 391, 288
9
Crameri et al., Nature, 1998, 391, 288
10
Crameri et al., Nature, 1998, 391, 288
11
but these dont bind competitively to the
receptor! Therefor the library was biased
AGAINST the WxxxGLW motif at the DNA level
Yanofsky et al., PNAS, 1996, 93, 7381
12
Yanofsky et al., PNAS, 1996, 93, 7381
13
Yanofsky et al., PNAS, 1996, 93, 7381
14
Cunningham et al., Science, 1991, 254, 821
15
Cunningham et al., Science, 1991, 254, 821
16
Cunningham et al., Science, 1991, 254, 821
17
Wells, PNAS, 1996, 93, 1
18
Wells, PNAS, 1996, 93, 1
19
Wells, PNAS, 1996, 93, 1
20
Li et al., Science, 1995, 270, 1657
21
Li et al., Science, 1995, 270, 1657
22
Li et al., Science, 1995, 270, 1657
23
Braisted and Wells, PNAS, 1996, 93, 5688
24
Braisted and Wells, PNAS, 1996, 93, 5688
Nedwidek M N , Hecht M H PNAS 1997 9410010
25
Braisted and Wells, PNAS, 1996, 93, 5688
26
Braisted and Wells, PNAS, 1996, 93, 5688
27
Braisted and Wells, PNAS, 1996, 93, 5688
28
Starovasnik et al, PNAS, 1997, 94, 10080
29
Starovasnik et al, PNAS, 1997, 94, 10080
30
Gao et al, PNAS, 2009, 106, 3071
31
Gao et al, PNAS, 2009, 106, 3071
32
Gao et al, PNAS, 2009, 106, 3071