1. A polypeptide fragment comprising a polypeptide of S2 ectodomain fragment
of spike protein of Coronavirus linked to a polypeptide of receptor binding domain
(RBD) fragment of spike protein of Coronavirus, at C-terminal or N-terminal end
5 of the S2 ectodomain fragment,
wherein the polypeptide of the S2 ectodomain fragment comprises a
polypeptide having at least 80%, at least 85%, at least 90%, at least 95%, at
least 98% or at least 99% sequence identity to a sequence selected from SEQ
ID. NO. 65 or SEQ ID. NO. 74,
10 wherein the sequence of the polypeptide of the S2 ectodomain fragment is
selected from the group consisting of
(a) SEQ ID. NO. 65 comprising amino acid substitution with proline at
amino acid positions corresponding to 120, 195, 202, 245, 289, and 290;
(b) SEQ ID. NO. 65 comprising amino acid substitution with proline at
15 amino acid positions corresponding to 120, 195, 202, and 245, and amino acid
substitution with aspartic acid at amino acid positions corresponding to 272
and 276;
(c) SEQ ID. NO. 65 comprising amino acid substitution with aspartic acid
at amino acid positions corresponding to 272 and 276;
20 (d) SEQ ID. NO. 74 comprising amino acid substitution with proline at
amino acid positions corresponding to 120, 195, 202, 245, 289, and 290;
(e) SEQ ID. NO. 74 comprising amino acid substitution with proline at
amino acid positions corresponding to 120, 195, 202, and 245, amino acid
substitution with aspartic acid at amino acid positions corresponding to 272
25 and 276; and
(f) SEQ ID. NO. 74 comprising amino acid substitution with aspartic acid
at amino acid positions corresponding to 272 and 276,
67
wherein the polypeptide of the receptor binding domain fragment comprises a
polypeptide having at least 80%, 85%, 90%, 95%, 98% or 99% sequence
identity to a sequence as set forth in SEQ ID. NO. 83 (RBD2) or SEQ ID. NO.
86 (RBD1),
5 wherein the sequence of the polypeptide of the receptor binding domain
(RBD) fragment is selected from the group consisting of
(g) SEQ ID. NO. 83 comprising amino acid substitution with proline at
amino acid positions corresponding to 17, amino acid substitution with
tryptophan at a amino acid position corresponding to 34 and amino acid
10 substitution with leucine at amino acid position corresponding to 196; and
(h) SEQ ID. NO. 86 comprising amino acid substitution with proline at
amino acid position corresponding to 18, amino acid substitution with
tryptophan at amino acid position corresponding to 35, and amino acid
substitution with leucine at amino acid position corresponding to 197.
15 2. The polypeptide fragment as claimed in claim 1, wherein the polypeptide of
receptor binding domain fragment is having at least one sequence selected from a
group consisting of SEQ ID. NO. 84 and SEQ ID. NO. 87.
3. The polypeptide fragment as claimed in claim 1 or 2, wherein the polypeptide
of S2 ectodomain fragment having the sequence selected from SEQ ID. NO. 65 or
20 SEQ ID. NO. 74 further comprises amino acid substitutions with hydrophilic amino
acids in at least three or at least four amino acid positions selected from the group
consisting of 158, 164, 167, 279, and 287.
4. The polypeptide fragment as claimed in claim 3, wherein the amino acid
substitution with hydrophilic amino acid is selected from a group consisting of
25 F158S, L164E, L167D, V279D, and L287Q.
5. The polypeptide fragment as claimed in any one of claims 1 to 4, wherein the
polypeptide of S2 ectodomain fragment is having at least one sequence selected
from the group consisting of SEQ ID NO. 66, SEQ ID NO. 68, SEQ ID NO. 75, and
SEQ ID NO. 77.
68
6. The polypeptide fragment as claimed in any one of claims 1 to 5, wherein the
N-terminal of the polypeptide of S2 ectodomain fragment is linked, by a linker
polypeptide, to C-terminal of the polypeptide of the receptor binding domain
fragment; or the C-terminal of the polypeptide of S2 ectodomain fragment is linked,
5 by a linker polypeptide, to N-terminal of the polypeptide of the receptor binding
domain fragment.
7. The polypeptide fragment as claimed in any one of claims 1 to 6, wherein the
polypeptide fragment is of a sequence selected from the group consisting of SEQ
ID. NO. 1, SEQ ID. NO. 5, SEQ ID. NO. 9, SEQ ID. NO. 13, SEQ ID. NO. 17, SEQ
10 ID. NO. 21, SEQ ID. NO. 25, SEQ ID. NO. 29, SEQ ID. NO. 33, SEQ ID. NO. 37,
SEQ ID. NO. 41, SEQ ID. NO.45, SEQ ID. NO. 49, SEQ ID. NO. 53, SEQ ID. NO.
57, and SEQ ID. NO. 61.
8. A polypeptide fragment comprising a polypeptide of S2 ectodomain fragment
having at least 80%, at least 85%, at least 90%, at least 95%, at least 98% or at least
15 99% sequence identity to a sequence selected from SEQ ID. NO. 65 or SEQ ID.
NO. 74,
wherein the sequence of the polypeptide of the S2 ectodomain fragment is selected
from the group consisting of
(a) SEQ ID. NO. 65 comprising amino acid substitution with proline at amino acid
20 positions corresponding to 120, 195, 202, 245, 289, and 290;
(b) SEQ ID. NO. 65 comprising amino acid substitution with proline at amino acid
positions corresponding to 120, 195, 202, and 245, and amino acid substitution with
aspartic acid at amino acid positions corresponding to 272 and 276;
(c) SEQ ID. NO. 65 comprising amino acid substitution with aspartic acid at amino
25 acid positions corresponding to 272 and 276;
(d) SEQ ID. NO. 74 comprising amino acid substitution with proline at amino acid
positions corresponding to 120, 195, 202, 245, 289, and 290;
69
(e) SEQ ID. NO. 74 comprising amino acid substitution with proline at amino acid
positions corresponding to 120, 195, 202, and 245, amino acid substitution with
aspartic acid at amino acid positions corresponding to 272 and 276; and
(f) SEQ ID. NO. 74 comprising amino acid substitution with aspartic acid at amino
5 acid positions corresponding to 272 and 276,
9. The polypeptide fragment as claimed in claim 8, wherein the polypeptide of S2
ectodomain fragment having the sequence selected from SEQ ID. NO. 65 or SEQ
ID. NO. 74 further comprises amino acid substitutions with hydrophilic amino acids
in at least three or at least four amino acid positions selected from the group
10 consisting of 158, 164, 167, 279, and 287.
10. The polypeptide fragment as claimed in claim 9, wherein the amino acid
substitution with hydrophilic amino acid is selected from a group consisting of
F158S, L164E, L167D, V279D, and L287Q.