Toxin gene from Xenorhabdus nematophilus

Details Toxin gene from Xenorhabdus nematophilus Toxin gene from Xenorhabdus nematophilus

1996-04-05

1999-10-26

Low, Christopher S. F.

Chemistry: molecular biology and microbiology

Micro-organism, per se ; compositions thereof; proces of...

Bacteria or actinomycetales; media therefor

4353201, 43525233, 43525411, 536 231, 536 232, 536 237, 800295, 800302, 800301, 935 9, 935 10, 935 22, 935 28, 935 31, 935 32, 935 38, 935 25, 935 64, 935 68, 935 72, C12N 121, C12N 115, C12N 701

Patent

active

059726873

DESCRIPTION:

BRIEF SUMMARY
TECHNICAL FIELD

The present invention concerns the identification and isolation of a new class of protein toxins specific against insects which are produced by bacteria from the species Xenorhabdus nematophilus and possibly by the species X.beddingii. In addition, the present invention relates to the insertion of this class of toxin into recombinant viruses, bacteria, protozoa, fungi, and transgenic plants in order to broaden the use of these toxins for control of a large range of insect pests and plant parasitic nematodes.


BACKGROUND

Insect pathogenic nematodes of the family Steinernematidae are known to be symbiotically associated with bacteria of the genus Xenorhabdus. It has been observed that these bacteria have the ability to kill a wide range of different insects without the aid of their nematode partners.
The present inventors have identified a new class of toxins. A DNA fragment encoding one of these toxins has been isolated from Xenorhabdus nematophilus stain A24 and characterised by sequencing. As will be recogised by persons skilled in the art, DNA fragments encoding members of this new class of toxins may be usefully introduced into viral agents, including entomopox and nuclear polyhedrosis viruses; bacteria (including Gracilicutes, Firmicutes, Tenericutes and Mendosicutes); fungi; protozoa; and plants.


SUMMARY OF THE PRESENT INVENTION

In a first aspect, the present invention consists in a polynucleotide molecule comprising a nucleotide sequence which encodes an insecticidal toxin and which is at least 70% homologous to the nucleotide sequence shown in Table 1 from residue 83 to 919 (SEQ ID NO:1), or a fragment thereof which fragment encodes an insecticidal polypeptide.
In a preferred embodiment of the present invention the nucleotide sequence is at least 90% homologous to the sequence shown in Table 1 from residue 83 to 919 (SEQ ID NO:1).
Preferably, the nucleotide sequence which encodes an insecticidal toxin from Xenorhabdus and more preferably, the nucleotide sequence substantially corresponds to the sequence shown in Table 1 from residue 83 to 919 (SEQ ID NO:1).
In a second aspect the present invention provides an insecticidal toxin which includes an amino acid sequence which is at least 70% homologous to residues 1 to 278 (SEQ ID NO:2) shown in Table 2 or a functional fragment thereof.
In a preferred embodiment of the present invention the insecticidal toxin includes an amino acid sequence which is at least 90% homologous to residues 1 to 278 (SEQ ID NO:2) shown in Table 2 or a functional fragment thereof.
In a further preferred embodiment the insecticidal toxin includes an amino acid sequence substantially corresponding to residues 1 to 278 (SEQ ID NO:2) in Table 2 or a functional fragment thereof.
In a third aspect the present invention provides a recombinant organism, the organism being characterised in that it is transformed with the polynucleotide molecule of the first aspect of the present invention.
The organisms which may be usefully transformed with the polynucleotide molecule of the first aspect of the present invention include viral agents such as entomopox and nuclear polyhedrosis viruses; bacteria, such as Gracilicutes, Firmicutes, Tenericutes and Mendosicutes; fungi; protozoa; and plants.
The term "substantially corresponds" as used herein in relation to the nucleotide sequence is intended to encompass minor variations in the nucleotide sequence which due to degeneracy do not result in a change in the encoded protein. Further this term is intended to encompass other minor variations in the sequence which may be required to enhance expression in a particular system but in which the variations do not result in a decrease in biological activity of the encoded protein.
The term "substantially corresponding" is used herein in relation to the amino acid sequence is intended to encompass minor variations in the amino acid sequence which do not result in a decrease in biological activity of the insecticidal toxin. These variations may include conservative amino acid substitut

REFERENCES:
Weiss (May 30, 1997) Washington Post, p. A01.
Balcerzak et al., 1991, Acta Parasitol. Pol. 36(4): 175-181, in: Biol. Abstr. 94(1): AB-610, abstract 5707.
Jarosz et al., 1991, Entomophaga 36(3): 361-368, in: Biol. Abstr. 93(1): AB-610, abstract 5688.
Gaugler et al., eds., 1990, in: Entomopathogenic Nematodes in Biolgoical Control, CRC Press, Boca Raton, FL. pp. 76, 77, and 85.
B. V. McInerney et al. "Biologically Active Metabolites from Xenorhabdus SPP., Part 1. Dithiolopyrrolone Derivatives with antibiotic Activity" Journal of Natural Products (1991) 54:774-784.
B. V. McInerney et al. "Biologically Active Metabolites from Xenorhabdus SPP., Part 2. Benzopyran-1-one Derivatives with Gastroprotective Activity" Journal of Natural Products (1991) 54:784-795.
M.A. Innis et al. "PCR Protocols: A Guide to methods and Applications" Chapter 11 "Cloning with PCR" by S. J. Scharf pp. 84-91.

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