Combinatorial libraries of monomer domains
Methods for identifying discrete monomer domains and immuno-domains with a desired property are provided. Methods for generating multimers from two or more selected discrete monomer domains are also provided, along with methods for identifying multimers possessing a desired property. Presentation systems are also provided which present the discrete monomer and/or immuno-domains, selected monomer and/or immuno-domains, multimers and/or selected multimers to allow their selection. Compositions, libraries and cells that express one or more library member, along with kits and integrated systems, are also included in the present invention.
1 - 92 . (canceled)
93 . A product comprising at least two monomer domains, wherein at least one monomer domain is a non-naturally occurring monomer domain and the monomer domains bind calcium.
94 . The product of claim 93 , wherein more than one of the monomer domains is a non-naturally-occurring monomer domain.
95 . The product of claim 93 , wherein each of the monomer domains is a non-naturally-occurring monomer domain.
96 . The product of claim 93 , wherein each of the monomer domains binds calcium.
97 . The product of claim 93 , wherein at least one of the monomer domains is derived from an LDL-receptor class A domain.
98 . The product of claim 93 , wherein at least one of the monomer domains is derived from an EGF-like domain.
99 . The product of claim 93 , wherein at least one domain has a binding specificity for a blood factor.
100 . The product of claim 99 , wherein the blood factor is serum albumin.
101 . The product of claim 93 , wherein the monomer domains are separated by a linker.
102 . The product of claim 101 , wherein the linker is a peptide linker.
103 . The product of claim 102 , wherein the linker is between 4 to 12 amino acids long.
104 . The product of claim 93 , wherein the product comprises a first monomer domain that binds a first molecule and a second monomer domain that binds a second molecule.
105 . The product of claim 104 , wherein the first and second molecules are different.
106 . The product of claim 104 , wherein the first and second molecules are different copies of the same molecule.
107 . The product of claim 93 , wherein the product comprises two monomer domains, each monomer domain having a binding specificity for a binding site on a first molecule.
108 . The product of claim 107 , wherein each of the two monomer domains have a binding specificity for a different binding site on the first molecule.
109 . The product of claim 93 , wherein the monomer domains are between 25 and 500 amino acids.
110 . The product of claim 93 , wherein the monomer domains are between 25 and 50 amino acids.
111 . The product of claim 93 , wherein the monomer domains are between 30 and 100 amino acids.
112 . The product of claim 93 , wherein the product comprises at least three monomer domains.
113 . The product of claim 93 , wherein the product comprises four monomer domains.
114 . The product of claim 93 , wherein the monomer domains are derived from human monomer domains.
115 . The product of claim 93 , wherein a polypeptide comprises the at least two monomer domains.
116 . A product comprising at least 4 monomer domains, wherein at least one monomer domain is non-naturally occurring, and wherein:
a. each monomer domain is between 30-100 amino acids and each of the monomer domains comprise at least one disulfide linkage; or
b. each monomer domain is between 30-100 amino acids and is derived from an extracellular protein; or
c. each monomer domain is between 30-100 amino acids and binds to a protein target.
117 . The product of claim 116 , wherein each monomer domain is between 30-100 amino acids and each of the monomer domains comprise at least one disulfide linkage.
118 . The product of claim 116 , wherein each monomer domain is between 30-100 amino acids and is derived from an extracellular protein.
119 . The product of claim 116 , wherein each monomer domain is between 30-100 amino acids and binds to a protein target.
120 . The product of claim 116 , wherein the monomer domains are derived from human monomer domains.
121 . The product of claim 116 , wherein more than one of the monomer domains is a non-naturally-occurring monomer domain.
122 . The product of claim 116 , wherein each of the monomer domains is a non-naturally-occurring monomer domain.
123 . The product of claim 116 , wherein each of the monomer domains binds a metal ion.
124 . The product of claim 116 , wherein each of the monomer domains binds a calcium ion.
125 . The product of claim 116 , wherein at least one of the monomer domains is derived from an LDL-receptor class A domain.
126 . The product of claim 116 , wherein at least one of the monomer domains is derived from an EGF-like domain.
127 . The product of claim 116 , wherein at least one domain has a binding specificity for a blood factor.
128 . The product of claim 127 , wherein the blood factor is serum albumin.
129 . The product of claim 116 , wherein the monomer domains are separated by a linker.
130 . The product of claim 129 , wherein the monomer domains are separated by a peptide linker.
131 . The product of claim 130 , wherein the linker is between 4 to 12 amino acids long.
132 . The product of claim 116 , wherein the product comprises a first monomer domain that binds a first molecule and a second monomer domain that binds a second molecule.
133 . The product of claim 132 , wherein the first and second molecules are different.
134 . The product of claim 133 , wherein the first and second molecules are different copies of the same molecule.
135 . The product of claim 116 , wherein the product comprises two monomer domains, each monomer domain having a binding specificity for a site on a first molecule.
136 . The product of claim 135 , wherein each of the two monomer domains have a binding specificity for a different binding site on the first molecule.
137 . The product of claim 116 , wherein a polypeptide comprises the at least four monomer domains.
138 . A product comprising at least 4 monomer domains, wherein at least one monomer domain is non-naturally occurring, and wherein:
a. each monomer domain is between 35-100 amino acids; or
b. each domain comprises at least one disulfide bond and is derived from a human protein and/or an extracellular protein.
139 . The product of claim 138 , wherein each monomer domain is between 35-100 amino acids.
140 . The product of claim 138 , wherein each domain comprises at least one disulfide bond and is derived from a human protein and/or an extracellular protein.
141 . The product of claim 138 , wherein the monomer domains are derived from human monomer domains.
142 . The product of claim 138 , wherein more than one of the monomer domains is a non-naturally-occurring monomer domain.
143 . The product of claim 138 , wherein each of the monomer domains is a non-naturally-occurring monomer domain.
144 . The product of claim 138 , wherein each of the monomer domains binds a metal ion.
145 . The product of claim 138 , wherein each of the monomer domains binds calcium.
146 . The product of claim 138 , wherein at least one of the monomer domains is derived from an LDL-receptor class A domain.
147 . The product of claim 138 , wherein at least one of the monomer domains is derived from an EGF-like domain.
148 . The product of claim 138 , wherein at least one domain has a binding specificity for a blood factor.
149 . The product of claim 148 , wherein the blood factor is serum albumin.
150 . The product of claim 138 , wherein the monomer domains are separated by a linker.
151 . The product of claim 150 , wherein the monomer domains are separated by a peptide linker.
152 . The product of claim 151 , wherein the linker is between 4 to 12 amino acids long.
153 . The product of claim 138 , wherein the product comprises a first monomer domain that binds a first molecule and a second monomer domain that binds a second molecule.
154 . The product of claim 153 , wherein the first and second molecules are different.
155 . The product of claim 154 , wherein the first and second molecules are different copies of the same molecule.
156 . The product of claim 138 , wherein the product comprises two monomer domains, each monomer domain having a binding specificity for a different site on a first molecule.
157 . The product of claim 157 , wherein each of the two monomer domains have a binding specificity for a different binding site on the first molecule.
158 . The product of claim 138 , wherein the product comprises at least three monomer domains.
159 . The product of claim 138 , wherein the product comprises four monomer domains.
160 . The product of claim 138 , wherein a polypeptide comprises the at least four monomer domains.
161 . A product comprising at least two monomer domains, wherein at least one monomer domain is non-naturally occurring, and wherein each domain is:
a. 25-50 amino acids long and comprises at least one disulfide bond; or
b. 25-50 amino acids long and is derived from an extracellular protein; or
c. 25-50 amino acids and binds to a protein target; or
d. 35-50 amino acids long.
162 . The product of claim 161 , wherein each domain is 25-50 amino acids long and comprises at least one disulfide bond.
163 . The product of claim 161 , wherein each domain is 25-50 amino acids and binds to a protein target.
164 . The product of claim 161 , wherein each domain is 25-50 amino acids long and is derived from an extracellular protein.
165 . The product of claim 161 , wherein each domain is 35-50 amino acids long.
166 . The product of claim 165 , wherein each domain is 35-45 amino acids long.
167 . The product of claim 161 , wherein more than one of the monomer domains is a non-naturally-occurring monomer domain.
168 . The product of claim 161 , wherein each of the monomer domains is a non-naturally-occurring monomer domain.
169 . The product of claim 161 , wherein each of the monomer domains binds calcium.
170 . The product of claim 161 , wherein at least one of the monomer domains is derived from an LDL-receptor class A domain.
171 . The product of claim 161 , wherein at least one of the monomer domains is derived from an EGF-like domain.
172 . The product of claim 161 , wherein at least one domain has a binding specificity for a blood factor.
173 . The product of claim 172 , wherein the blood factor is serum albumin.
174 . The product of claim 161 , wherein the monomer domains are separated by a linker.
175 . The product of claim 174 , wherein the monomer domains are separated by a peptide linker.
176 . The product of claim 175 , wherein the linker is between 4 to 12 amino acids long.
177 . The product of claim 161 , wherein the product comprises a first monomer domain that binds a first molecule and a second monomer domain that binds a second molecule.
178 . The product of claim 177 , wherein the first and second molecules are different.
179 . The product of claim 178 , wherein the first and second molecules are different copies of the same molecule.
180 . The product of claim 161 , wherein the product comprises two monomer domains, each monomer domain having a binding specificity for a different site on a first molecule.
181 . The product of claim 156 , wherein each of the two monomer domains have a binding specificity for a different binding site on the first molecule.
182 . The product of claim 161 , wherein the product comprises at least three monomer domains.
183 . The product of claim 161 , wherein the product comprises four monomer domains.
184 . The product of claim 161 , wherein a polypeptide comprises the at least two monomer domains.
185 . A product comprising at least two monomer domains, wherein at least one monomer domain is non-naturally-occurring and each monomer domain comprises at least two disulfide bonds.
186 . The product of claim 185 , wherein each monomer domain comprises at least three disulfide bonds.
187 . The product of claim 185 , wherein at least one monomer domain is derived from an extracellular protein.
188 . The product of claim 185 , wherein at least one monomer domain binds to a target protein.
189 . The product of claim 185 , wherein more than one of the monomer domains is a non-naturally-occurring monomer domain.
190 . The product of claim 185 , wherein each of the monomer domains is a non-naturally-occurring monomer domain.
191 . The product of claim 185 , wherein each of the monomer domains binds calcium.
192 . The product of claim 185 , wherein at least one of the monomer domains is derived from an LDL-receptor class A domain.
193 . The product of claim 185 , wherein at least one of the monomer domains is derived from an EGF-like domain.
194 . The product of claim 185 , wherein at least one domain has a binding specificity for a blood factor.
195 . The product of claim 194 , wherein the blood factor is serum albumin.
196 . The product of claim 185 , wherein the monomer domains are separated by a linker.
197 . The product of claim 196 , wherein the monomer domains are separated by a peptide linker.
198 . The product of claim 197 , wherein the linker is between 4 to 12 amino acids long.
199 . The product of claim 185 , wherein the product comprises a first monomer domain that binds a first molecule and a second monomer domain that binds a second molecule.
200 . The product of claim 199 , wherein the first and second molecules are different.
201 . The product of claim 200 , wherein the first and second molecules are different copies of the same molecule.
202 . The product of claim 185 , wherein the product comprises two monomer domains, each monomer domain having a binding specificity for a different site on a first molecule.
203 . The product of claim 202 , wherein each of the two monomer domains have a binding specificity for a different binding site on the first molecule.
204 . The product of claim 185 , wherein the product comprises at least three monomer domains.
205 . The product of claim 185 , wherein the product comprises four monomer domains.
206 . The product of claim 185 , wherein the product comprises the at least two monomer domains.
207 . A method for identifying a monomer domain with affinity for a molecule, the method comprising,
providing a library of monomer domains, wherein each monomer domain:
is between 30-100 amino acids;
comprises at least one disulfide bond; and
binds an ion; and
screening the library of monomer domains for affinity to a first molecule; and
identifying at least one monomer domain that binds to at least one molecule.
208 . The method of claim 207 , wherein the ion is calcium.
209 . The method of claim 207 , further comprising,
screening the library of monomer domains for affinity to a second molecule;
identifying a monomer domain that binds to a second molecule;
linking at least one monomer domain with affinity for the first molecule with at least one monomer domain with affinity for the second molecule, thereby forming a multimer with affinity for the first and the second molecules.
210 . The method of claim 207 , further comprising,
linking the identified monomer domain to a library of monomer domains to form a library of multimers, each multimer comprising at least two monomer domains;
screening the library of multimers for the ability to bind to the first molecule or a second molecule; and
identifying a multimer that binds to the first molecule or second molecule.
211 . The method of claim 210 , wherein the library of multimers is screened for the ability to bind to the first molecule.
212 . The method of claim 210 , wherein the library of multimers is screened for the ability to bind to the second molecule.
213 . A method for identifying a multimer that binds to at least one molecule, the method comprising:
providing a library of multimers, wherein each multimer comprises at least two monomer domains, and wherein at least one monomer:
is between 30-100 amino acids;
comprises a t least one disulfide bond; and
binds an ion; and
screening the library of multimers for molecule-binding multimers.
214 . The method of claim 213 , wherein the ion is calcium.
215 . A library of multimers, wherein the multimers comprise at least two monomer domains connected by a linker; and the monomer domains
are between 30-100 amino acids;
comprise a t least one disulfide bond; and
bind an ion.
216 . The library of claim 215 , wherein the ion is calcium.