Albumin variants and conjugates

1. A conjugation-competent polypeptide comprising an amino acid sequence which is at least 90% identical to a human albumin having the sequence set forth in SEQ ID NO: 2;

wherein at least one position equivalent to a position selected from K93, A226, E230, I271, E294, E358, L24, F49, V54, D56, A92, Q94, E97, H128, F156, E227, D237, K240, D259, K262, N267, Q268, L275, L284, K317, A322, E333, D340, E354, K359, A362, E382, or L398 of said human albumin having the sequence set forth in SEQ ID NO: 2 comprises a substitution to a cysteine such that said conjugation-competent polypeptide comprises one or more conjugation-competent cysteine residues; or

wherein said conjugation-competent polypeptide comprises an insertion of a cysteine at a position adjacent to an amino acid corresponding to a position equivalent to any of residues K93, A226, E230, I271, E294, E358, L24, F49, V54, D56, L66, A92, Q94, E97, H128, F156, E227, D237, K240, D259, K262, N267, Q268, L275, E277, L284, E311, K317, A322, E333, D340, E354, K359, A362, E382, or L398 of said human albumin having the sequence set forth in SEQ ID NO: 2.

2. The conjugation-competent polypeptide of claim 1 , wherein said conjugation-competent polypeptide comprises conjugation-competent cysteine residues at two, three, four, five, six, or seven positions selected from K93, A226, E230, I271, E294, E358, L24, F49, V54, D56, L66, A92, Q94, E97, H128, F156, E227, D237, K240, D259, K262, N267, Q268, L275, E277, L284, E311, K317, A322, E333, D340, E354, K359, A362, E382, or L398 of said human albumin having the sequence set forth in SEQ ID NO: 2.

3. The conjugation-competent polypeptide of claim 1 , wherein said conjugation-competent polypeptide lacks a conjugation-competent cysteine at a position equivalent to position 34 of said human albumin having the sequence set forth in SEQ ID NO: 2.

4. The conjugation-competent polypeptide of claim 1 , comprising two or more conjugation-competent cysteine residues, wherein when the polypeptide is folded, there is a distance of at least 5 Å between at least two of the conjugation-competent cysteine residues, as determined by X-ray crystallography or Nuclear Magnetic Resonance Spectroscopy.

5. The conjugation-competent polypeptide of claim 1 , wherein said conjugation-competent polypeptide comprises a substitution of a cysteine at one or both positions corresponding to residues K93 or E294 of said human albumin having the sequence set forth in SEQ ID NO: 2.

6. The conjugation-competent polypeptide of claim 1 , wherein said conjugation-competent polypeptide is configured to form a conjugate with maleimide-polyethylenglycol2-biotin at a conjugation efficiency of at least 95% and, wherein said conjugate is at least 95% stable in aqueous solution.

7. A conjugation-competent polypeptide comprising an amino acid sequence, which is at least 90% identical to a human albumin having the sequence set forth in SEQ ID NO: 2;

wherein at least one position equivalent to a position selected from K93, A226, E230, I1271, E294, E358, L24, F49, V54, D56, L66, A92, Q94, E97, H128, F156, E227, D237, K240, D259, K262, N267, Q268, L275, E277, L284, E311, K317, A322, E333, D340, E354, K359, A362, E382, or L398 of said human albumin having the sequence set forth in SEQ ID NO: 2 is substituted with a cysteine and, wherein said conjugation-competent polypeptide further comprises:

a substitution or insertion of a cysteine at or adjacent to position D1, A2, H3, S5, A55, S58, C75, T76, T79, E82, T83, E86, C91, D121, V122, C124, T125, D129, C169, C177, A229, T236, E266, D269, S270, S273, S304, K313, D314, C316, N318, A320, C361, A364, C369, A371, N386, Q390, Q397, S435, T478, T496, A504, E505, T506, T508, D549, C558, D562, C567, A581, L585 or A578 of said human albumin having the sequence set forth in SEQ ID NO. 2; or

a deletion or substitution of a cysteine at a position corresponding to any of C360, C316, C75, C168, C558, C361, C91, C124, C169 or C567 of said human albumin having the sequence set forth in SEQ ID NO: 2 so as to generate a conjugation competent cysteine at any of C369, C361, C91, C177, C567, C316, C75, C169, C124 or C558; or

addition of a cysteine to the N-side of the N-terminal residue of an albumin sequence or to the C-side of the C-terminal residue of an albumin sequence; or

one or more substitutions, which alter the binding affinity of the polypeptide for FcRn; or one or more substitutions, which alter the plasma half-life of the polypeptide.

8. The conjugation-competent polypeptide of claim 7 , wherein said conjugation-competent polypeptide comprises conjugation-competent cysteines at: (a) A2+L585, (b) A2+A364+D562+L585, (c) A2 and adjacent the C-side of the C-terminus (d) T79+A364; (e) A364+D1; (f) T79+D562+A364; (g) D562+A364+D1; (h) T79+D562+A364+A504; (i) T79+D562+A364+L585; (j) T79+D562+A364+D1; (k) T79+D562+A364+L585+D1; (1) E86+D562+A364+A504+A2; (m) S270+A581; (n) S270+D129; (o) S270+A581+E82; (p) S270+A581+D129; (q) S270+A581+E82+D129; (r) S270+A581+E82+D129+Q397; (s) C369+C177; (t) A364+A581; (u) T79+A364+A581; (v) A364+A581+D129; (w) A364+C177; (x) D562+C369; (y) D129+C369; (z) A581+C369; or (aa) D562+D129+C369.

9. The conjugation-competent polypeptide of claim 7 , wherein the one or more substitutions which alters the binding affinity of the polypeptide for FcRn, or alters the plasma half-life of the polypeptide is at one or more positions in said human albumin having the sequence set forth in SEQ ID NO: 2 selected from: 573, 500, 550, 417, 440, 464, 490, 492, 493, 494, 495, 496, 499, 501, 503, 504, 505, 506, 510, 535, 536, 537, 538, 540, 541, 542, 574, 575, 577, 578, 579, 580, 581, 582 or 584.

10. The conjugation-competent polypeptide of claim 7 , wherein the one or more substitutions, which alters the binding affinity of the polypeptide for FcRn, or alters the plasma half-life of the polypeptide corresponds to: K573Y, W, P, H, F, V, I, T, N, S, G, M, C, A, E, Q, R, L, D, K500E, G, D, A, S, C, P, H, F, N, W, T, M, Y, V, Q, L, I, R, Q417A, H440A, H464Q, E492G, D494N,Q,A, E495Q,A, T496A, D494E+Q417H, D494N+T496A, E492G+V493P, P499A, E501A,Q, N503H,K, H510Q, H535Q, K536A, P537A, K538A, K541G,D, D550E,N, E492G+K573P,A, or E492G/N503H/K573P of said human albumin having the sequence set forth in SEQ ID NO: 2.

11. The conjugation-competent polypeptide of claim 7 , wherein the polypeptide comprises alterations at two or more positions selected (a) 492 and 580; (b) 492 and 574; (c) 492 and 550; (d) 550 and 573; (e) 550 and 574; or (f) 550 and 580 of said human albumin having the sequence set forth in SEQ ID NO: 2.

12. The conjugation-competent polypeptide of claim 1 comprising one or more substitutions in Domain I of said human albumin having the sequence set forth in SEQ ID NO: 2; and one or more substitutions in Domain III of said human albumin having the sequence set forth in SEQ ID NO: 2, wherein said conjugation-competent polypeptide has an altered binding affinity to FcRn.

13. The conjugation-competent polypeptide of claim 12 , wherein the substitutions in Domain I are selected from any of positions 78 to 120 of said human albumin having the sequence set forth in SEQ ID NO: 2 and the substitutions in Domain III are selected from any of positions 425, 505, 510, 512, 524, 527, 531, 534, 569, 573, or 575 of said human albumin having the sequence set forth in SEQ ID NO: 2.

14. The conjugation-competent polypeptide of claim 12 , wherein the substitutions in Domain I and Domain III are selected from (i) 83N, K or S; (ii) 111D, G, H, R, Q or E; or (iii) 573P, Y, W, H, F, T, I or V.

15. The conjugation-competent polypeptide of claim 1 comprising one or more substitutions in Domain II of said human albumin having the sequence set forth in SEQ ID NO: 2 selected from positions 349, 342, 381, 345, 384, 198, 206, 340, 341, 343, 344, 352, 382, 348, or 383 of said human albumin having the sequence set forth in SEQ ID NO: 2; wherein the one or more substitutions causes said conjugation-competent polypeptides to have (i) an altered plasma half-life or (ii) an altered binding affinity to FcRn.

16. The conjugation-competent polypeptide of claim 15 , wherein the substitution at position 349, 342, 381, 345, 384, 198, 206, 340, 341, 343, 344, 352, 382, 348, or 383 is selected from (i) 349F, W, Y, H, P, K or Q; (ii) 342Y, W, F, H, T, N, Q, A, C, I, L, P, V; (iii) 381G or A; or (iv) 345E, H, I or Q.

17. The conjugation-competent polypeptide of claim 15 , comprising one or more substitutions in said human albumin having the sequence set forth in SEQ ID NO: 2 selected from positions V418, T420, V424, E505, V547, or K573 of said human albumin having the sequence set forth in SEQ ID NO: 2; wherein the one or more substitutions causes the conjugation-competent polypeptides to have (i) an altered plasma half-life or (ii) an altered binding affinity to FcRn.

18. The conjugation-competent polypeptide of claim 1 , further comprising one or more substitutions in said human albumin having the sequence set forth in SEQ ID NO: 2 selected from V381, E383, N391, Y40, K402, L407, Y411, K413, K414, V415C, Q416, V424, V426D, G434, E442, R445, P447, E450, S454, V455, V456, L457, Q459, L463, E495, T506, T508, F509, A511, D512, T515, L516, S517, K519, R521, I523, K524, K525, Q526, T527, E531, H535, K538, A539, K541, K557, A561, T566, or A569, of said human albumin having the sequence set forth in SEQ ID NO: 2; wherein the one or more substitutions causes the conjugation-competent polypeptides to have (i) an altered plasma half-life or (ii) an altered binding affinity to FcRn.

19. The conjugation-competent polypeptide of claim 1 comprising one or more substitutions in said human albumin having the sequence set forth in SEQ ID NO: 2 selected from V547, K573, I523, T527, K500, or E505 of said human albumin having the sequence set forth in SEQ ID NO: 2; wherein the one or more substitutions causes the conjugation-competent polypeptides to have (i) an altered plasma half-life or (ii) an altered binding affinity to FcRn.

20. The conjugation-competent polypeptide of claim 1 comprising one or more substitutions in said human albumin having the sequence set forth in SEQ ID NO: 2 selected from positions 573, 523, 527 or 505 of said human albumin having the sequence set forth in SEQ ID NO: 2.

21. The conjugation-competent polypeptide of claim 1 , wherein the propensity of said conjugation-competent polypeptide to exist as a monomer in solution is enhanced relative to the propensity of the polypeptide of SEQ ID NO. 2 to exist as a monomer in solution; is equal to the propensity of the polypeptide of SEQ ID NO. 2 to exist as a monomer in solution; or is reduced by no more that 30% relative to the propensity of the polypeptide of SEQ ID NO. 2 to exist as a monomer in solution.

22. A method of increasing the half-life of a molecule by fusing or conjugating said molecule to a polypeptide of claim 1 , or by associating said molecule with a polypeptide of claim 1 , wherein said molecule comprises a bioactive agent, an imaging agent, a diagnostic agent, a contrast agent or a therapeutic compound.

23. A method of preparing a conjugation-competent polypeptide, the method comprising:

providing a nucleic acid encoding a conjugation-competent polypeptide which is at least 90% identical to a human albumin having the sequence set forth in SEQ ID NO: 2, wherein at least one position equivalent to a position selected from K93, A226, E230, I271, E294, E358, L24, F49, V54, D56, A92, Q94, E97, H128, F156, E227, D237, K240, D259, K262, N267, Q268, L275, L284, K317, A322, E333, D340, E354, K359, A362, E382, or L398 of said human albumin having the sequence set forth in SEQ ID NO: 2 comprises a substitution to a cysteine such that said conjugation-competent polypeptide comprises one or more conjugation-competent cysteine residues; or

wherein said conjugation-competent polypeptide comprises an insertion of a cysteine at a position adjacent to an amino acid corresponding to a position equivalent to any of residues K93, A226, E230, I271, E294, E358, L24, F49, V54, D56, L66, A92, Q94, E97, H128, F156, E227, D237, K240, D259, K262, N267, Q268, L275, E277, L284, E311, K317, A322, E333, D340, E354, K359, A362, E382, or L398 of said human albumin having the sequence set forth in SEQ ID NO: 2; and

expressing the protein product of said nucleic acid to yield said conjugation-competent polypeptide.

24. The conjugation-competent polypeptide of claim 5 , wherein said conjugation-competent polypeptide comprises a substitution of a cysteine at position corresponding to residue K93 of said human albumin having the sequence set forth in SEQ ID NO: 2.

25. The conjugation-competent polypeptide of claim 5 , wherein said conjugation-competent polypeptide comprises a substitution of a cysteine at position corresponding to residue E294 of said human albumin having the sequence set forth in SEQ ID NO: 2.