Skip to main content

Table 1 Rationale for CDR design

From: Structure-based design and construction of a synthetic phage display nanobody library

Sequence position Original aa Degenerate codon Coded aa Analysis
CDR1
 26 Gly Gly Conserved (our own analysis). An outlier in the Ramachandran plot. Keep fixed
 27 Gly DMY Ala:2 Asn:2 Asp:2
Ser:2 Thr:2 Tyr:2
An outlier in the Ramachandran plot. Randomize, but keep it small. The degenerate codon includes also Tyr
 28 Ser VRN Arg:6 Asn:2 Asp:2
Gln:2 Glu:2 Gly:4
His:2 Lys:2 Ser:2
Surface-exposed side chain. Set it variable, but polar
 29 Glu
 30 Tyr HWY Asn:2 His:2 Ile:2
Leu:2 Phe:2 Tyr:2
It is packed against the extremes of CDR1. Keep it mostly big and hydrophobic/aromatic
 31 Ser VRN see above Surface-exposed side chain. Set it variable, but polar
 32 Tyr Tyr Buried aa playing an important packing role. Keep fixed
 33 Ser DMY see above It’s placed on an alpha turn, outside the binding site. Keep it mostly small, as the original aa
 34 Thr VNN Ala:4 Arg:6 Asn:2 Asp:2 Gln:2 Glu:2 Gly:4 His:2 Ile:3 Leu:4 Lys:2 Met:1 Pro:4 Ser:2 Thr:4 Val:4 Points “upwards” at the base of CDR1. Set it very variable
 35 Phe Phe Buried aa playing an important packing role. Keep fixed
 36 Ser KMY Ala:2 Asp:2 Ser:2 Tyr:2 Support for CDR H3 (found also making a Cys bridge with CDR H3). Becomes buried by long CDRs 3. Small aa are commonly found, as well as Tyr
CDR2
 53 Ala Ala Within a β-sheet. Packed with the long CDR3. Keep fixed
 54 Ile Ile Very conserved, buried aa playing an important packing role. Keep fixed
 55 Ala KMY see above Interacting with the bent CDR3 in the original Nb. Ser and Tyr are commonly found, as well as small aa
 56 Ser WBG Arg:1 Leu:1 Met:1
Ser:1 Thr:1 Trp:1
Small aa side chains, like Ser and Thr, remain partially buried. The larger amino acids in the set would pack against CDR1 and the N-ter of CDR3
 57 Met VRN see above Surface-exposed side chain. Set it variable, but polar
 58, 59 Gly Gly Conserved pair of residues, outliers in the Ramachandran plot. Keep fixed
 60 Leu ADM Arg:1 Asn:1 Ile:2
Lys:1 Ser:1
Combines long charged aa. that can protrude upwards with small polar aa. Arg, Asn Ile and Ser are among the most frequently found aa at this position
CDR3
 VNN VRN WMY VRN WMY VNN WMY VRN WMY VNN WMY TWY VRN TMY WMY: Asn:2 Ser:2 Thr:2 Tyr:2
TWY: Phe:2 Tyr:2
TMY: Ser:2 Tyr:2
IUPAC degenerate base symbols [14]:           
B = C, G, T H = A, C, T M = A, C R = A, G V = A, C, G Y = C, T D = A, G, T K = G, T N = A, C, G, T S = C, G W = A, T
  1. Numbers in bold correspond to CDR positions that were randomized