Plant-produced recombinant aprotinin and aprotinin variants
OwnerFakhrieh Vojdani, Kenneth Palmer +2inventors
InventorsFakhrieh Vojdani, Kenneth Palmer, Stephen Garger, Gregory Pogue
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Abstract
The present invention relates to plant produced native aprotinin and aprotinin variants having enzyme-inhibitory, immunological and pharmacokinetic properties and their preparation. In a preferred method a recombinant RNA plant virus is used to express native aprotinin+variants thereof in Nicotiana plants.
Claims22
A composition comprising a plant produced recombinant variant aprotinin, wherein the variant aprotinin has an amino acid other than methionine at position 52.
An isolated DNA molecule, comprising
- d. a DNA sequence encoding a variant aprotinin, wherein the variant aprotinin has an amino acid other than methionine at position 52;
- and
- e. a DNA sequence encoding an RNA subgenomic promoter;
- f. wherein the DNA sequence encoding aprotinin having a oxidation-resistant amino acid at position 52 is attached at its 5′ end to the DNA sequence encoding an RNA subgenomic promoter so that when a resulting negative sense RNA molecule encoded by the DNA molecule is present, then expression of the aprotinin-encoding DNA sequence is allowed in a plant.
An isolated RNA molecule, comprising:
- d. a plus sense single stranded RNA subgenomic promoter sequence, and
- e. a plus sense single stranded RNA sequence encoding a variant aprotinin, wherein the variant aprotinin has an amino acid other than methionine at position 52;
- f. wherein the sequence encoding the variant aprotinin is linked at its 5′ end to said plus sense single stranded RNA subgenomic promoter sequence.
A recombinant single stranded plus sense plant viral RNA, comprising:
- f. an RNA coding sequence for 126-kDa and 183-kDa replicase subunits;
- g. a first coat protein subgenomic promoter sequence being attached at its 3′ end to an RNA sequence coding for a variant aprotinin, wherein the variant aprotinin has an amino acid other than methionine at position 52.
A recombinant single stranded plus sense plant viral RNA, further comprising:
- h. the RNA coding sequence for 126-kDa and 183-kDa replicase subunits attached at its 3′ end to a 30-kDa open reading frame of a viral movement protein,
- i. the 30-kDa open reading frame of a viral movement protein containing the first coat protein subgenomic promoter sequence;
- j. the RNA sequence coding for variant aprotinin attached at its 3′ end to a second coat protein subgenomic promoter sequence;
- k. the second coat protein subgenomic promoter sequence attached at its 3′ end to a coat protein coding sequence.
A plant-compatible expression vector comprising an artificial polynucleotide encoding a recombinant variant aprotinin, wherein the variant aprotinin has an amino acid other than methionine at position 52.
Dependencies are inferred from the claim text.