Proteins toxic to hemipteran insect species
The present invention discloses Hemipteran insect inhibitory proteins, methods of using such proteins, nucleotide sequences encoding such proteins, methods of detecting and isolating such proteins, and their use in agricultural systems.
1. A recombinant DNA molecule comprising a polynucleotide encoding a polypeptide comprising a sequence having at least 95% amino acid identity to SEQ ID NO: 2 (TIC807), and wherein said polypeptide comprises:
a) at least one modification at an amino acid residue selected from the group consisting of Val10, Asn12, Ile14, Asn22, Asn23, Gly24, Ile25, Gln26, Gly27, Phe30, Gln38, Ile39, Asp40, Thr41, Ile43, Ile52, Thr93, Ser95, Ser97, Phe138, Phe147, Gln149, Ser151, Ser159, Asn180, Thr182, Ser193, Thr194, Glu195, His196, Tyr197, Ser198, His199, Tyr200, Ser201, Gly202, Tyr203, Pro204, Ile205, Leu206, Thr207, Trp208, Ile209, Ser210, Tyr216, Gly218, Pro219, Met221, Ser222, Trp223, Tyr224, Phe225, Asn239, Va1244, Tyr246, Thr247, Va1251, Gln253, Ser255, and His287 relative to SEQ ID NO:2; and
(b) a contiguous triple deletion in residue range 196-201 relative to SEQ ID NO:2;
wherein said polypeptide exhibits about 15 fold or greater Hemipteran inhibitory activity against a Hemipteran pest species relative to the Hemipteran inhibitory activity of SEQ ID NO: 2.
2. The polypeptide of claim 1 , wherein said polypeptide exhibits about 25 fold or greater Hemipteran inhibitory activity against a Hemipteran pest species relative to the Hemipteran inhibitory activity of SEQ ID NO: 2 (TIC807).
3. The polypeptide of claim 1 , wherein said polypeptide comprises at least one mutation in SEQ ID NO:2 selected from the group consisting of N12D, I52M, T93A, S95A, F138V, F147S, F147A, Q149E, Q149A, S151A, S159T, N180D, T182A, H196A, Y197A, S198A, W208A, P219R, W223Y, Y246F, T247K, V251A, and H287F.
4. The polypeptide of claim 1 , wherein said polypeptide comprises at least two mutations in SEQ ID NO:2 selected from the group consisting of T93A, S95A, F147S, F147A, Q149E, Q149A, S151A, N180D, T182A, H196A, Y197A, S198A, W208A, S217N, P219R, W223Y, V251A.
5. The polypeptide of claim 1 , wherein said polypeptide comprises at least three mutations in SEQ ID NO:2 selected from the group consisting of T93A, S95A, F147S, F147A, Q149E, Q149A, S151A, N180D, T182A, H196A, S198A, Y197A, W208A, S217N, P219R, W223Y, V251A.
6. The polypeptide of claim 1 , wherein said polypeptide comprises at least four mutations in SEQ ID NO:2 selected from the group consisting of T93A, S95A, F147S, F147A, Q149E, Q149A, S151A, N180D, T182A, H196A, Y197A, S198A, W208A, S217N, P219R, W223Y, V251A.
7. A host cell comprising the polynucleotide of claim 1 , wherein the host cell is selected from the group consisting of a bacterial host cell and a plant host cell.
8. The host cell of claim 7 , wherein the bacterial host cell is selected from the group consisting of Agrobacterium, Rhizobium, Bacillus, Escherichia, Pseudomonas , and Salmonella.
9. The host cell of claim 7 , wherein the plant host cell is selected from the group of consisting of a monocot cell and a dicot cell.
10. An insect inhibitory composition comprising the polypeptide of claim 2 .
11. The composition of claim 10 , wherein the composition is prepared by lyophilization, extraction, filtration, or centrifugation.
12. The insect inhibitory composition of claim 10 , further comprising at least one insect inhibitory agent different from the insect inhibitory recombinant polypeptide.
13. The insect inhibitory composition of claim 12 , wherein the at least one insect inhibitory agent is selected from the group consisting of an insect inhibitory protein, an insect inhibitory dsRNA molecule, and an insect inhibitory chemistry.
14. The insect inhibitory composition of claim 12 , wherein the at least one pesticidal agent exhibits activity against one or more pest species of the order Lepidoptera, Coleoptera, Hemiptera, or Homoptera.
15. A seed comprising the polynucleotide of claim 1 .
16. A transgenic plant cell, plant, or plant part comprising the polynucleotide of claim 1 .
17. A method of controlling a Hemipteran pest, comprising exposing the Hemipteran pest to the transgenic plant cell, plant, or plant part of claim 16 , wherein the plant cell, plant, or plant part expresses a Hemipteran inhibitory amount of the polypeptide encoded by said polynucleotide.
18. A commodity product obtained from the plant cell, plant, or plant part of claim 16 , wherein the commodity product comprises a detectable amount of the polypeptide encoded by said polynucleotide, wherein the commodity product is selected from the group consisting of plant biomass, oil, meal, animal feed, flour, flakes, bran, lint, hulls, and processed seed.
19. A method of producing a seed, the method comprising:
planting at least one seed comprising said polynucleotide of claim 1 ;
growing at least one plant from the at least one seed; and
harvesting a seed from the at least one plant, wherein the harvested seed comprises said polynucleotide.