Enhancing spinosyn production with oxygen binding proteins

Abstract

The invention describes the integration of polynucleotides into chromosomal DNA of S. spinosa species, which are useful for the production of insecticides, integrants thereof, and also to the use of the integrants. The invention includes the stable integration and expression of an oxygen-binding protein, VHb, which results in increased spinosyn production.

What is claimed is: 
     
         1 . A method for increasing a spinosyn titer comprising:
 intergrating an oxygen binding protein into a spinosyn producing strain, wherein the spinosyn titer is increased by at least 4.6%.   
     
     
         2 . The method of  claim 1  wherein the oxygen binding protein is a globin protein. 
     
     
         3 . The method of  claim 2  wherein the globin protein is selected from the group consisting of  Vitreoscilla  hemoglobin,  Alcaligenes eutrophus  flavohemoprotein, horse heart myoglobin,  E. coli  hemoprotein,  B. subtilis  hemoprotein, yeast flavohemoglobin, soybean leghemoglobin, lupin leghemoglobin, and sperm whale myoglobin, or their functional equivalents. 
     
     
         4 . The method of  claim 2  wherein the globin protein is  Vitreoscilla  hemoglobin or a functional polymorphism thereof. 
     
     
         5 . The method of  claim 1  wherein the oxygen binding protein is integrated into chromosomal DNA in a neutral site in a  S. spinosa  genome. 
     
     
         6 . The method of  claim 1  wherein the spinosyn titers are increased are selected from the group consisting of spinsoyn A+D, spinosyn J+L, spinosyn A and spinosyn J. 
     
     
         7 . A method of increasing spinosyn production, the method comprising:
 culturing a spinosyn producing organism that expresses a heterologous gene encoding an oxygen binding protein under conditions appropriate for production of spinosyn; and   collecting spinosyn from a culture of an spinosyn producing organism that expresses a heterologous gene encoding an oxygen binding protein.   
     
     
         8 . The method of  claim 7  wherein the organism is  S. spinosa.    
     
     
         9 . The method of  claim 7  wherein the heterologous gene is integrated into a chromosome of the cell. 
     
     
         10 . The method of  claim 7  wherein the oxygen binding protein is a globin protein. 
     
     
         11 . The method of  claim 10  wherein the globin protein is selected from the group consisting of  Vitreoscilla  hemoglobin,  Alcaligenes eutrophus  flavohemoprotein, horse heart myoglobin,  E. coli  hemoprotein,  B. subtilis  hemoprotein, yeast flavohemoglobin, soybean leghemoglobin, lupin leghemoglobin, and sperm whale myoglobin, or their functional equivalents. 
     
     
         12 . The method of  claim 10  wherein the globin protein is  Vitreoscilla  hemoglobin or a functional polymorphism thereof. 
     
     
         13 . The method of  claim 7  wherein the oxygen binding protein is integrated into chromosomal DNA in a neutral site in a  S. spinosa  genome.