SYNTHETIC POLYPEPTIDES AND USES THEREOF

1 . A synthetic polypeptide comprising or consisting of an amino acid sequence with at least 85% sequence identity to one of the following sequences or functional fragments thereof:

Syn2-N

(SEQ ID NO: 425)

CLSYDTEILTVEYGLIPIGEIVEKKIECTVYTIDNNGLIYTQSIEQWHHR

GYQELFEYILEDGSTIRATKDHKFMTSERQMLPIEEIFERGWELKQVL;

Syn3-N

(SEQ ID NO: 426)

CLSSDTEVITEEYGPIAIGKIVDEGIRCSVYSVDNNGNLYTQPISQWHDR

GRQEIYEYYLENGSVIRATKDHKFMTKDGEMLPIDEIFEKGLELKQVLP;

Syn5-N

(SEQ ID NO: 427)

CLSYETEVLTVEYGFMPIGKIVEERIRCSVYTVDKNGFIYSQPIAQWHQR

GLQEVYEYDLENGSIIRATKEHQFMTNDGQMLAIHEIFTRKLDLLQSQE;

Syn1-C

(SEQ ID NO: 428)

MKVISRKSLGTQPVYDICVTHDHNFLMKNGLIASN;

Syn4-C

(SEQ ID NO: 429)

MDVKIVSYKFLGSENVYDILERDHNFLIKNGLVASN;

Syn5-C

(SEQ ID NO: 430)

MVKIITYKSLGRQKVYDLGLEQDHNFVLANGLVASN;

Syn9-C

(SEQ ID NO: 431)

MVKIISRKYLDTQPVYDVGVQKDHNFLISNGSIASN;

and

Syn10-C

(SEQ ID NO: 432)

MVKIATRRSLGTEPVYDIGLQQEHNFLLANGLVASN.

2 . The synthetic polypeptide of claim 1 , wherein the synthetic polypeptide comprises an intein.

3 . A polynucleotide encoding the synthetic polypeptide of claim 1 .

4 . A cell comprising the polynucleotide of claim 3 .

5 . A pair of vectors, wherein:

a) one member of the pair of vectors comprises a polynucleotide sequence encoding a synthetic polypeptide-N(Syn-N) of claim 1 ; and

b) the other member of the pair of vectors comprises a polynucleotide sequence encoding a synthetic polypeptide-C(Syn-C) with at least about 85% amino acid sequence identity to a sequence selected from the group consisting of:

Syn1-C

(SEQ ID NO: 428)

MKVISRKSLGTQPVYDICVTHDHNFLMKNGLIASN;

Syn4-C

(SEQ ID NO: 429)

MDVKIVSYKFLGSENVYDILERDHNFLIKNGLVASN;

Syn5-C

(SEQ ID NO: 430)

MVKIITYKSLGRQKVYDLGLEQDHNFVLANGLVASN;

Syn9-C

(SEQ ID NO: 431)

MVKIISRKYLDTQPVYDVGVQKDHNFLISNGSIASN;

and

Syn10-C

(SEQ ID NO: 432)

MVKIATRRSLGTEPVYDIGLQQEHNFLLANGLVASN.

6 . The pair of vectors of claim 5 , wherein each vector is a PAL family AAV vector comprising a VP1 capsid polypeptide comprising an amino acid sequence with at least 95% amino acid sequence identity to the following AAV9 VP1 capsid polypeptide amino acid sequence with a 7-mer peptide inserted between amino acid positions Q588 and A589 relative to the following AAV9 VP1 capsid polypeptide amino acid sequence:

(SEQ ID NO: 443)

MAADGYLPDWLEDNLSEGIREWWALKPGAPQPKANQQHQDNARGLVLPGY

KYLGPGNGLDKGEPVNAADAAALEHDKAYDQQLKAGDNPYLKYNHADAEF

QERLKEDTSFGGNLGRAVFQAKKRLLEPLGLVEEAAKTAPGKKRPVEQSP

QEPDSSAGIGKSGAQPAKKRLNFGQTGDTESVPDPQPIGEPPAAPSGVGS

LTMASGGGAPVADNNEGADGVGSSSGNWHCDSQWLGDRVITTSTRTWALP

TYNNHLYKQISNSTSGGSSNDNAYFGYSTPWGYFDFNRFHCHFSPRDWQR

LINNNWGFRPKRLNFKLFNIQVKEVTDNNGVKTIANNLTSTVQVFTDSDY

QLPYVLGSAHEGCLPPFPADVFMIPQYGYLTLNDGSQAVGRSSFYCLEYF

PSQMLRTGNNFQFSYEFENVPFHSSYAHSQSLDRLMNPLIDQYLYYLSKT

INGSGQNQQTLKFSVAGPSNMAVQGRNYIPGPSYRQQRVSTTVTQNNNSE

FAWPGASSWALNGRNSLMNPGPAMASHKEGEDRFFPLSGSLIFGKQGTGR

DNVDADKVMITNEEEIKTTNPVATESYGQVATNHQSA QA QAQTGWVQNQG

ILPGMVWQDRDVYLQGPIWAKIPHTDGNFHPSPLMGGFGMKHPPPQILIK

NTPVPADPPTAFNKDKLNSFITQYSTGQVSVEIEWELQKENSKRWNPEIQ

YTSNYYKSNNVEFAVNTEGVYSEPRPIGTRYLTRNL;

and wherein the 7-mer peptide is selected from those listed in Table 7B.

7 . The pair of vectors of claim 6 , wherein the AAV vectors comprise the amino acid alterations A587D and Q588G relative to the AAV9 VP1 capsid polypeptide sequence.

8 . The pair of vectors of claim 5 , wherein the Syn-N and the Syn-C are each fused to a heterologous polypeptide.

9 . The pair of vectors of claim 8 , wherein the Syn-N is an N-intein and the Syn-C is a C-intein, which together are capable of functioning in protein splicing.

10 . A cell comprising the pair of vectors of claim 6 .

11 . A fusion protein comprising or consisting of a heterologous polypeptide fragment fused at the C-terminus thereof to a synthetic polypeptide of claim 1 .

12 . A fusion protein comprising a heterologous polypeptide fused at the N-terminus thereof to a synthetic polypeptide of claim 1 .

13 . A polynucleotide encoding the fusion protein of claim 12 .

14 . A vector comprising the polynucleotide of claim 13 .

15 . A cell comprising the vector of claim 14 .

16 . A pharmaceutical composition comprising the fusion protein of claim 12 and a pharmaceutically acceptable excipient.

17 . A polynucleotide delivery system comprising:

(a) a first polynucleotide encoding a fusion protein comprising a heterologous polypeptide fused at the C-terminus thereof to a first synthetic polypeptide of claim 1 ; and

(b) a second polynucleotide encoding a fusion protein comprising another heterologous polypeptide fused at the N-terminus thereof to a second synthetic polypeptide, wherein the second synthetic polypeptide comprises an amino acid sequence with at least 85% sequence identity to one of the following sequences, or functional fragments thereof:

Syn1-C

(SEQ ID NO: 428)

MKVISRKSLGTQPVYDICVTHDHNFLMKNGLIASN;

Syn4-C

(SEQ ID NO: 429)

MDVKIVSYKFLGSENVYDILERDHNFLIKNGLVASN;

Syn5-C

(SEQ ID NO: 430)

MVKIITYKSLGRQKVYDLGLEQDHNFVLANGLVASN;

Syn9-C

(SEQ ID NO: 431)

MVKIISRKYLDTQPVYDVGVQKDHNFLISNGSIASN;

and

Syn10-C

(SEQ ID NO: 432)

MVKIATRRSLGTEPVYDIGLQQEHNFLLANGLVASN.

18 . A polynucleotide delivery system comprising:

(a) a first polynucleotide encoding a fusion protein comprising the N-terminal fragment of a base editor, wherein the base editor comprises a deaminase domain, and a nucleic acid programmable DNA binding protein (napDNAbp) domain, and a first synthetic polypeptide fused to the C-terminus of the N-terminal fragment of the base editor, wherein the first synthetic polypeptide is a polypeptide of claim 1 ; and

(b) a second polynucleotide encoding a fusion protein comprising a second synthetic polypeptide fused to the N-terminus of the C-terminal fragment of the base editor, wherein the second synthetic polypeptide comprises an amino acid sequence with at least 85% sequence identity to one of the following sequences, or functional fragments thereof:

Syn1-C

(SEQ ID NO: 428)

MKVISRKSLGTQPVYDICVTHDHNFLMKNGLIASN;

Syn4-C

(SEQ ID NO: 429)

MDVKIVSYKFLGSENVYDILERDHNFLIKNGLVASN;

Syn5-C

(SEQ ID NO: 430)

MVKIITYKSLGRQKVYDLGLEQDHNFVLANGLVASN;

Syn9-C

(SEQ ID NO: 431)

MVKIISRKYLDTQPVYDVGVQKDHNFLISNGSIASN;

and

Syn10-C

(SEQ ID NO: 432)

MVKIATRRSLGTEPVYDIGLQQEHNFLLANGLVASN.

19 . A method of delivering polynucleotides encoding heterologous polypeptides to a cell, the method comprising contacting the cell with:

(a) a first polynucleotide encoding a fusion protein comprising a heterologous polypeptide or a fragment thereof fused at the C-terminus thereof to a first synthetic polypeptide of claim 1 ; and

(b) a second polynucleotide encoding a fusion protein comprising another heterologous polypeptide fused at the N-terminus thereof to a second synthetic polypeptide, wherein the second synthetic polypeptide comprises an amino acid sequence with at least 85% sequence identity to one of the following sequences, or functional fragments thereof:

Syn1-C

(SEQ ID NO: 428)

MKVISRKSLGTQPVYDICVTHDHNFLMKNGLIASN;

Syn4-C

(SEQ ID NO: 429)

MDVKIVSYKFLGSENVYDILERDHNFLIKNGLVASN;

Syn5-C

(SEQ ID NO: 430)

MVKIITYKSLGRQKVYDLGLEQDHNFVLANGLVASN;

Syn9-C

(SEQ ID NO: 431)

MVKIISRKYLDTQPVYDVGVQKDHNFLISNGSIASN;

and

Syn10-C

(SEQ ID NO: 432)

MVKIATRRSLGTEPVYDIGLQQEHNFLLANGLVASN.

20 . A method for delivering polynucleotides encoding base editor fragments to a cell, the method comprising contacting the cell with:

(a) a first polynucleotide encoding a fusion protein comprising the N-terminal fragment of a base editor, where the base editor comprises a deaminase domain, and a nucleic acid programmable DNA binding protein (napDNAbp) domain, and a first synthetic polypeptide fused to the C-terminus of the N-terminal fragment of the base editor, wherein the first synthetic polypeptide is a polypeptide of claim 1 ; and

(b) a second polynucleotide encoding a fusion protein comprising a second synthetic polypeptide fused to the N-terminus of the C-terminal fragment of the base editor, wherein the second synthetic polypeptide comprises an amino acid sequence with at least 85% sequence identity to one of the following sequences, or functional fragments thereof:

Syn1-C

(SEQ ID NO: 428)

MKVISRKSLGTQPVYDICVTHDHNFLMKNGLIASN;

Syn4-C

(SEQ ID NO: 429)

MDVKIVSYKFLGSENVYDILERDHNFLIKNGLVASN;

Syn5-C

(SEQ ID NO: 430)

MVKIITYKSLGRQKVYDLGLEQDHNFVLANGLVASN;

Syn9-C

(SEQ ID NO: 431)

MVKIISRKYLDTQPVYDVGVQKDHNFLISNGSIASN;

and

Syn10-C

(SEQ ID NO: 432)

MVKIATRRSLGTEPVYDIGLQQEHNFLLANGLVASN.

21 . A method for editing a target polynucleotide in a cell, the method comprising delivering polynucleotides encoding base editor fragments to a cell according to the method of claim 20 .

22 . A kit suitable for use in the method of claim 21 .