Combinatorial Libraries Based on C-type Lectin-like Domain
AssigneeAnaphore, Inc.
InventorsMartha Wild, Anke Kretz-Rommel, Katherine S. Bowdish, Mark Renshaw
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Abstract
This invention relates to polypeptide libraries comprising polypeptides having a C-type lectin domain (CTLD) with a randomized loop region, as well as nucleic acid libraries comprising nucleic acid molecules encoding such polypeptides. The invention also relates to methods for generating the randomized polypeptides and the polypeptide libraries. The invention further relates to methods of screening the polypeptide and nucleic acid libraries based on the specific binding of the modified CTLDs to a target molecule of interest. The invention also relates to polypeptides derived from such libraries that bind to target molecules of interest.
Claims41
A combinatorial polypeptide library comprising polypeptide members that comprise a C-type lectin domain (CTLD) having a randomized loop region, wherein the CTLD loop region comprises loop segment A (LSA) containing Loops 1-4 and loop segment B (LSB) containing Loop 5 and is randomized according to one of the following Schemes:
- (a) amino acid modifications in at least one of the four loops in loop segment A (LSA) of the CTLD, wherein the amino acid modifications comprise an insertion of at least one amino acid in Loop 1 and random substitution of at least five amino acids within Loop 1;
- (b) amino acid modifications in at least one of the four loops in loop segment A (LSA) of the CTLD, wherein the amino acid modifications comprise random substitution of at least five amino acids within Loop 1 and random substitution of at least three amino acids within Loop 2;
- (c) amino acid modifications in at least one of the four loops in the loop segment A (LSA) of the CTLD, wherein the amino acid modifications comprise random substitution of at least seven amino acids within Loop 1 and at least one amino acid insertion in Loop 4;
- (d) amino acid modifications in at least one of the four loops in the loop segment A (LSA) of the CTLD, wherein the amino acid modifications comprise at least one amino acid insertion in Loop 3 and random substitution of at least three amino acids within Loop 3;
- (e) amino acid modifications in at least one of the four loops in the loop segment A (LSA) of the CTLD, wherein the amino acid modifications comprise a modification that combines two loops into a single loop, wherein the two combined loops are Loop 3 and Loop 4;
- (f) amino acid modifications in at least one of the four loops in the loop segment A (LSA) of the CTLD, wherein the amino acid modifications comprise at least one amino acid insertion in Loop 4 and random substitution of at least three amino acids within Loop 4;
- (g) amino acid modifications in at least one of the four loops in the loop segment A (LSA) of the CTLD and in loop segment B (LSB), wherein the amino acid modifications comprise random substitution of at least five amino acid residues in Loop 3 and random substitution of at least three amino acids within Loop 5;
- (h) amino acid modifications in at least one of the four loops in the loop segment A (LSA) of the CTLD, wherein the amino acid modifications comprise random substitution of at least one amino acid and insertion of at least six amino acids in Loop 3;
- (i) amino acid modifications in at least one of the four loops in the loop segment A (LSA) of the CTLD, wherein the amino acid modifications comprise a mixture of (1) random substitution of at least six amino acids in Loop 3 and (2) random substitution of at least six amino acids and at least one amino acid insertion in Loop 3;
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- (j) amino acid modifications in at least one of the four loops in the loop segment A (LSA) of the CTLD, wherein the amino acid modifications comprise at least four or more amino acid insertions in at least one of the four loops in the loop segment A (LSA) or loop 5 in loop segment B (LSB) of the CTLD.
Dependencies are inferred from the claim text.