Variants of native growth hormones having non naturally occurring amino acid sequences or covalent modifications
AssigneeGenentech, Inc.
InventorsBrian C. Cunningham, James A. Wells
Life of this patent
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Abstract
Growth hormones contain two receptor binding sites termed site 1 and site 2 with which they form ternary complexes with their receptors. In this scheme, site 1 on the hormone binds to one molecule of receptor and then site 2 binds to another molecule of receptor, thereby producing a 1:2 complex. Disclosed herein are growth hormone variants consisting of modifications to the native sequence in the region of site 1 or site 2, or alternatively a combination of modifications at both site 1 and site 2.
Claims25
A variant of a native mature mammalian growth hormone, said variant having a non-naturally occurring growth hormone amino acid sequence or a covalent modification, comprising a mutation selected from the group consisting of:
- (a) an amino acid substitution at a residue corresponding to residue 1 of human growth hormone, numbered from the N-terminus;
- (b) an amino acid substitution at a residue corresponding to residue 4 of human growth hormone, numbered from the N-terminus, wherein the substituted amino acid is selected from the group consisting of R, N, D, C, Q, E, G, H, K, M, F, P, S, T, W, Y, and V;
- (c) an amino acid substitution at a residue corresponding to residue of 6 of human growth hormone, numbered from the N-terminus, wherein the substituted amino acid is selected from the group consisting of R, N, D, C, Q, E, G, H, I, K, M, F, P, S, T, W, Y, and V;
- (d) an amino acid substitution at a residue corresponding to residue 8 of human growth hormone, numbered from the N-terminus, wherein the substituted amino acid is selected from the group consisting of N, D, C, Q, E, G, H, I, L, K, F, P, S, T, W, Y, M, and V;
- (e) insertion of an amino acid adjacent to a residue corresponding to a residue of human growth hormone selected from the group consisting of 1, 4, 6, and 8, numbered from the N-terminus;
- and
- (f) covalent modification of a residue corresponding to a residue of human growth hormone selected from the group consisting of 1, 4, 6, and 8, numbered from the N-terminus.
A variant of a native mature mammalian growth hormone, said variant having a non-naturally occurring amino acid sequence or a covalent modification, comprising:
- (a) a first mutation in a first region corresponding to residue 1 to residue 15 of human growth hormone, numbered from the N-terminus;
- and
- (b) a second mutation in a second region corresponding to residue 106 to residue 128 of human growth hormone, numbered from the N-terminus.
A variant of a native mature mammalian growth hormone, said variant having a non-naturally occurring amino acid sequence or a covalent modification, comprising:
- (a) a first mutation in a first region selected from the group consisting of (i) amino acid residue L15, R16, R19 and Q22, (ii) the C-terminal ⅔ of the loop linking helices 1 and 2, and (iii) the C-terminal ½ of helix 4;
- and
- (b) a second mutation in a second region selected from the group consisting of (i) a region corresponding to residue 1 to residue 15 of human growth hormone and (ii) a region corresponding to residue 106 to residue 128 of human growth hormone, numbered from the N-terminus.
A variant of a native, mature mammalian monomeric polypeptide ligand having four amphipathic alpha helices, wherein said native ligand binds to a first receptor polypeptide through a first ligand site and then binds to a second receptor polypeptide through a second ligand site, thereby forming a ternary complex, said variant having a non-naturally occurring amino acid sequence or a covalent modification and comprising:
- (a) a first mutation in a portion of said second ligand site corresponding to the first third of helix 1;
- and
- (b) a second mutation in a portion of a said second ligand site corresponding to the middle half of helix 3.
A variant of a native, mature mammalian monomeric polypeptide ligand having four amphipathic alpha helices, wherein said native ligand binds to a first receptor polypeptide through a first ligand site and then binds to a second receptor polypeptide through a second ligand site, thereby forming a ternary complex, said variant having a non-naturally occurring ammo acid sequence or a covalent modification and comprising:
- (a) a first mutation including an amino acid substitution in a portion of said first ligand site selected from the group consisting of (i) amino acid residue L15, R16, R19 and Q22, (ii) the C-terminal ⅔ of the loop linking helices 1 and 2, and (iii) the C-terminal ½ of helix 4;
- and
- (b) a second mutation in a portion of said second ligand site selected from the group consisting of (i) the first third helix 1 and (ii) the middle half of helix 3.
Dependencies are inferred from the claim text.