Method for the design of fibrillar collagen-mimetic peptide self-assemblies
Novel synthetic human scale collagen triple helices assemblies are disclosed. Methods of making self-assembling collagen mimetic peptides that self-assemble into human scale collagen are also disclosed.
1. A method for making a collagen mimetic peptide capable of self-assembly comprising:
(a) combining a plurality of self-assembling collagen mimetic peptides consisting essentially of Xaa-Yaa-Gly tripeptide repeats, wherein Xaa and Yaa are amino acids, wherein the number of tripeptide repeats (nT) is 3v±1, wherein v is a positive non-zero integer, wherein the peptide is n residues in length, wherein n=nT×3; and
(b) incubating the plurality of self-assembling collagen mimetic peptides of step (a) for a sufficient time to allow the self-assembling collagen mimetic peptides to self-assemble into a triple helical collagen assembly to form a composition,
wherein neighboring self-assembling collagen mimetic peptides of the triple helical collagen assembly are offset by v Xaa-Yaa-Gly tripeptide repeats,
wherein the Yaa amino acid of a K t Xaa-Yaa-Gly tripeptide repeat comprises an amino acid capable of charge pairing and the Xaa amino acid of a D t Xaa-Yaa-Gly tripeptide repeat comprises an amino acid capable of charge pairing with the Yaa amino acid of the Kt Xaa-Yaa-Gly tripeptide repeat on a neighboring self-assembling collagen mimetic peptide of the triple helical collagen assembly, and
wherein K t is a positive integer from 1 to nT and D t =(K t ±v+1) mod nT.
2. The method of claim 1 wherein assembly of the collagen mimetic peptides segregates a charged block at an end of the triple helical collagen assembly.
3. The method of claim 2 wherein the triple helical collagen assembly is stabilized by an interstrand interaction between the Yaa amino acid of the K t Xaa-Yaa-Gly tripeptide repeat and the Xaa amino acid of a D t Xaa-Yaa-Gly tripeptide repeat on the neighboring self-assembling collagen mimetic peptide of the triple helical collagen assembly.
4. The method of claim 3 , wherein the interstrand interaction comprises at least one interstrand salt bridge.
5. The method of claim 4 , wherein the at least one interstrand salt bridge is formed from a lysine residue in the Yaa position of the K t Xaa-Yaa-Gly tripeptide repeatand anaspartic acid residue in the Xaa position of the D t Xaa-Yaa-Gly tripeptide repeat on the neighboring self-assembling collagen mimetic peptide.
6. The method of claim 4 , wherein the at least one interstand salt bridge comprises a first interstrand salt bridge and a second interstrand salt bridge, wherein the first interstrand salt bridge is formed from a lysine residue in the Yaa position of the K t Xaa-Yaa-Gly tripeptide repeat on a first strand and an aspartic acid residue in the Xaa position of the D t Xaa-Yaa-Gly tripeptide repeat on the neighboring self-assembling collagen mimetic peptide on a second strand and the second interstrand salt bridge is formed from a lysine residue in the Yaa position of the K t Xaa-Yaa-Gly tripeptide repeat on a second strand and an aspartic acid residue in the Xaa position of the D t Xaa-Yaa-Gly tripeptide repeat on the neighboring self-assembling collagen mimetic peptide on a third strand.
7. The method of claim 4 , wherein the at least one interstrand salt bridge is formed from a lysine residue in the Yaa position on a first strand and an aspartic acid residue in the Xaa position on a second strand at a three residue offset.
8. The method of claim 2 further comprising combining the segregated charged block of a first multimer with a second collagen mimetic peptide or second multimer.
9. The method of claim 2 , further comprising combining a first mutltimer with a second collagen mimetic peptide or second multimer, wherein an end of the first multimer joins to a segregated charged block end of the second collagen mimetic peptide or second multimer.
10. The method of claim 1 wherein a plurality of collagen mimetic peptides assemble symmetrically to form a collagen assembly.
11. The method of claim 5 , wherein greater than about 90% of the available lysine residues and available aspartic acid residues are linked by an interstrand salt bridge.
12. The method of claim 1 , wherein nT is 3v+1.
13. The method of claim 1 , wherein nT is 3v−1.
14. The method of claim 1 , wherein v is 5.
15. The method of claim 14 , wherein the triple helical collagen assembly comprises a homotrimer wherein the homotrimer comprises about 4 salt bridges per about 14 residues.
16. The method of claim 1 , wherein the Xaa-Yaa-Gly tripeptide repeats comprises Xaa-Yaa-Gly tripeptide repeats of ProLysGly, AspHypGly, ProHypGly or combinations thereof.
17. The method of claim 1 , wherein the Xaa-Yaa-Gly tripeptide repeats are selected from the group consisting of POG, PKG, DOG, DKG and combinations thereof.
18. The method of claim 1 , wherein the plurality of collagen mimetic peptides assemble into a symmetrical triple helix wherein each strand of the helix comprises at least about 500 residues.
19. The method of claim 1 , wherein the plurality of collagen mimetic peptides assemble into a symmetrical triple helix of about 200 nm in length.
20. The method of claim 1 , wherein the self-assembling collagen mimetic peptide comprises a central Xaa-Yaa-Gly tripeptide repeat and Xaa of the central Xaa-Yaa-Gly tripeptide repeat is proline or aspartic acid and Yaa of the central Xaa-Yaa-Gly tripeptide repeat is lysine to reduce blunt end association.
21. The method of claim 1 , wherein the self-assembling collagen mimetic peptide comprises a terminal Xaa-Yaa-Gly tripeptide repeatand Yaa of the terminal Xaa-Yaa-Gly tripeptide repeat is hydroxyproline to reduce blunt end association.
22. The method of claim 1 , wherein the symmetrical triple helical collagen assembly has a melting temperature (Tm) of greater than about 37° C.
23. The method of claim 1 , wherein the symmetrical triple helical collagen assembly has a Tm of greater than about 45° C.
24. The method of claim 1 , wherein the Xaa-Yaa-Gly tripeptide repeats comprises any one of sequence SEQ ID NO: 2-9, 10, 12, 14-18, 22, or 27-37.
25. The method of claim 1 , wherein the collagen mimetic peptides comprise the tripeptides in the following sequence:
(POG)n(PKG/DOG)m(POG/DKG)p (DOG/PKG)q(POG)r wherein n, m, p, q, and r are selected from 0 or a positive integer of 1-10, and wherein nT=n+m+p+q+r=3v±1>0.