Granted Patent B2

US 12,378,539 · App. 18/915,656 · Granted Aug 5, 2025

Inventors: Jesse B. Owens (Honolulu, HI); Brian Hew (Honolulu, HI)

Assignee: University of Hawaii

C12N9/22

Abstract

Claims (108)

1. A Bxb1 integrase comprising:

an amino acid sequence that is at least 98% identical to the amino acid sequence of SEQ ID NO: 1; and

one or more amino acid mutations at positions selected from L4I, E42K, R57K, R63K, V76I, H111P, V122M, and V187I, relative to SEQ ID NO: 1.

2. The Bxb1 integrase of claim 1 , comprising an amino acid sequence that is at least 99% identical to the amino acid sequence of SEQ ID NO: 1.

3. The Bxb1 integrase of claim 1 , further comprising one or more amino acid mutations selected from D36N, V40A, A49S, V74I, I87L or I87A or I87S, H89G, V175A, V179A, R287H, A288V, T453I, and H496N.

4. The Bxb1 integrase of claim 1 , further comprising one or more amino acid mutations selected from V40I, D45G, I75V, H95Y, A119S, A280T, A311V, E434G, and V466M.

5. The Bxb1 integrase of claim 1 , wherein one or more of the following amino acids relative to SEQ ID NO: 1 are not mutated: A62, H100, G209, P295, L302, C307, L387, E419, S428, E483, and R487.

6. The Bxb1 integrase of claim 1 , comprising the mutation V122M.

7. The Bxb1 integrase of claim 6 , further comprising the mutation A369P.

8. A Bxb1 integrase comprising an amino acid sequence that is at least 99% identical to the amino acid sequence of SEQ ID NO: 1 and having the amino acid mutations I87L, A369P, and E434G; the amino acid mutations V122M, A369P, and E434G; or the amino acid mutations H95Y, A369P, and E434G.

9. A Bxb1 integrase comprising:

an amino acid sequence that is at least 98% identical to the amino acid sequence of SEQ ID NO: 1; and

one or more amino acid mutations selected from R319K, A341D, D359A, A369P, A398S, R416K, A425T, L479I, and M499T, relative to SEQ ID NO: 1.

10. The Bxb1 integrase of claim 9 , comprising an amino acid that is at least 99% identical to the amino acid sequence of SEQ ID NO: 1.

11. The Bxb1 integrase of claim 9 , further comprising one or more amino acid mutations selected from D36N, V40A, A49S, V74I, I87L or I87A or I87S, H89G, V175A, V179A, R287H, A288V, T453I, and H496N.

12. The Bxb1 integrase of claim 9 , further comprising one or more amino acid mutations selected from V40I, D45G, I75V, H95Y, A119S, A280T, A311V, E434G, and V466M.

13. The Bxb1 integrase of claim 9 , wherein one or more of the following amino acids relative to SEQ ID NO: 1 are not mutated: A62, H100, Q191, P195, G209, P295, L302, C307, L387, E419, S428, E483, and R487.

14. The Bxb1 integrase of claim 9 , further comprising the mutation V122M.

15. The Bxb1 integrase of claim 9 , comprising the mutation A369P.

16. A Bxb1 integrase comprising:

an amino acid sequence that is at least 98% identical to the amino acid sequence of SEQ ID NO: 1; and

a combination of amino acid mutations selected from:

A119S, A369P, and E434G;

A288V, A311V, A398S, R416K, T453I, H496N, and E42K,

A369P and E434G;

A49S, A369P, and E434G;

A49S, I87L, A369P, and E434G;

A49S, V122M, A369P, and E434G;

D36N, A369P, and E434G;

D36N, I87L, A369P, and E434G;

D36N, V122M, A369P, and E434G;

D45G, A369P, and E434G;

E42K, H95Y, A369P, and E434G;

E42K, V76I, and I87L;

H111P and E434G

H95, V122M and A369P;

H95Y and A341D;

H95Y and A369P;

H95Y and K313R;

H95Y and V122M;

H95Y, A369P, A425T, and E434G;

H95Y, A369P, and E434G;

H95Y, A369P, E434G, and L479I;

H95Y, A369P, E434G, and V466M;

H95Y, R319K, A369P, and E434G;

H95Y, V122M and E434G;

H95Y, V179A, A369P, and E434G;

I75V and V76I;

I87L and A341D;

I87L and A369P;

I87L and A425T;

I 87L and D359A;

I87L and L479I;

I87L and L4I;

I87L and R319K;

I87L and V122M;

I87L, A341D, A369P, and E434G;

I87L, A341D, and R287H;

I87L, A369P, A425T, and E434G;

I87L, A369P, and E434G;

I87L, A369P, E434G, and R319K;

I87L, D359A, A369P, and E434G;

I87L, H95Y, A369P and E434G;

I87L, H95Y, and A369P;

I87L, H95Y, V122M, and E434G;

I87L, R287H, A369P, and E434G;

I87L, R287H, A369P, E434G, and A341D;

I87L, R57K, and R63K;

I87L, V122, A369P, and E434G;

I87L, V122M, and A369P;

I87L, V122M, and E434G;

I87L, V175A, and D359A;

L4I and V76I;

L4I, H95Y, A369P, and E434G;

L4I, I87L, A369P, and E434G;

L4I, V122M, A369P, and E434G;

R57K, I87L, A369P, and E434G;

R57K, R63K and H95Y;

R57K, R63K, I87L, A369P, and E434G;

R57K, R63K, V122M, A369P, and E434G;

R63K and I87L;

R63K and V122M;

R63K, A119S, and M499T;

R63K, I87L, A369P, and E434G;

R63K, I87V, and N194D;

R63K, V122M, A369P, and E434G;

R63K, V122M, R319K, and V40I;

V122M and A369P;

V122M and E434G;

V122M, A369P, A425T, and E434G;

V122M, A369P, and E434G;

V122M, A369P, E434G, and A341D;

V122M, A369P, E434G, and R319K;

V122M, A369P, E434G, and V175A;

V122M, D359A, A369P, and E434G;

V122M, R287H, A341D, A369P, and E434G;

V122M, R287H, A369P, and E434G;

V122M, V175A, D359A, A369P, and E434G;

V40A, R63K, and H89G;

V40I, H95Y, A369P, and E434G;

V74I and V76I;

V74I, I75V, and V76I;

V76I and A119S;

V76I and A369P;

V76I and V122M;

V76I, A369P, and E434G;

V76I, I87L, and K313R; and

V76I, V122M, and A369P.

Assignments (1)

Continuity (4)

References Cited (124)