Process for production of avermectins and cultures therefor

Claim 2

Streptomyces avermitilis lacking one or both of the following activities: branched-chain 2-oxo-acid dehydrogenase, branched-chain amino acid transaminase.

Claim 3

Streptomyces avermitilis lacking branched-­chain 2-oxo acid dehydrogenase activity characterized by the inability of permeabilized cells thereof to produce ¹⁴CO₂ from added [¹⁴C-1]-2-oxoisocaproic acid.

Claim 4

Streptomyces avermitilis lacking branched-­chain 2-oxo acid dehydrogenase activity characterized by the inability of permeabilized cells thereof to produce ¹⁴CO₂ from added [¹⁴C-1]-2-oxoisocaproic acid.

Claim 5

Streptomyces avermitilis having the identify­ing characteristics of ATCC 53567 or ATCC 53568.

Claim 6

Streptomyces avermitilis having the identify­ing characteristics of ATCC 53567 or ATCC 53568.

Claim 7

Streptomyces avermitilis ATCC 53567 or ATCC 53568.

Claim 8

Streptomyces avermitilis ATCC 53567 or ATCC 53568.

Claim 9

Streptomyces avermitilis lacking branched-­chain amino acid transaminase characterized by the inability to grow on media lacking L-isoleucine, L-leucine and L-valine.

Claim 10

Streptomyces avermitilis lacking branched-­chain amino acid transaminase characterized by the inability to grow on media lacking L-isoleucine, L-leucine and L-valine.

Claim 11

Streptomyces avermitilis having the identify­ing characteristics of ATCC 53669 or ATCC 53670.

Claim 12

Streptomyces avermitilis having the identify­ing characteristics of ATCC 53669 or ATCC 53670.

Claim 13

Streptomyces avermitilis ATCC 53669 or ATCC 53670.

Claim 14

Streptomyces avermitilis ATCC 53669 or ATCC 53670.

Claim 15

Streptomyces avermitilis substantially incapable of producing substance C-076 upon fermenta­tion in an aqueous nutrient medium comprising an assimilable source of carbon, nitrogen and inorganic salts under aerobic conditions, said medium being substantially free of an acid of the formula R-COOH , or a compound convertible to said acid by S . avermitilis , wherein R is isopropyl or (S)-sec-butyl, but being capable of producing C-076 upon fermentation in said medium when said medium contains an acid of the formula RCOOH, or a compound convertible to said acid by S . avermitilis , wherein R is isopropyl or (S)-sec-­butyl.

Claim 16

Streptomyces avermitilis substantially incapable of producing substance C-076 upon fermenta­tion in an aqueous nutrient medium comprising an assimilable source of carbon, nitrogen and inorganic salts under aerobic conditions, said medium being substantially free of an acid of the formula R-COOH , or a compound convertible to said acid by S . avermitilis , wherein R is isopropyl or (S)-sec-butyl, but being capable of producing C-076 upon fermentation in said medium when said medium contains an acid of the formula RCOOH, or a compound convertible to said acid by S . avermitilis , wherein R is isopropyl or (S)-sec-­butyl.

Claim 17

Streptomyces avermitilis according to claim 8 having the identifying characteristics of ATCC 53567, ATCC 53568, ATCC 53669 or ATCC 53670.

Claim 18

Streptomyces avermitilis according to claim 9, said S . avermitilis being S . avermitilis ATCC 53567, ATCC 53568, ATCC 53669 or ATCC 53670.

Claim 19

A process for preparation of an avermectin which comprises aerobically fermenting with a strain of Streptomyces avermitilis which lacks one or both of branched-chain 2-oxo acid dehydrogenase activity and branched-chain amino acid transaminase activity, an aqueous nutrient medium comprising an assimilable source of nitrogen, carbon and inorganic salts and a compound capable of utilization in the biosynthesis of an avermectin.

Claim 20

A process for preparation of an avermectin which comprises aerobically fermenting with a strain of Streptomyces avermitilis which lacks one or both of branched-chain 2-oxo acid dehydrogenase activity and branched-chain amino acid transaminase activity, an aqueous nutrient medium comprising an assimilable source of nitrogen, carbon and inorganic salts and a compound capable of utilization in the biosynthesis of an avermectin.

Claim 21

A process according to claim 11 wherein the compound capable of utilization in the biosynthesis of an avermectin is an acid of the formula R-COOH or a compound convertible to said acid by S . avermitilis , wherein R is other than isopropyl or (S)-sec-butyl.

Claim 22

A process according to claim 11, wherein the S . avermitilis has the identifying characteristics of ATCC 53567, ATCC 53568, ATCC 53669 or ATCC 53670.

Claim 23

A process according to claim 11 wherein the compound is of the formula R-COOH wherein R is an alpha-branched-chain group, the carbon atom thereof to which is attached the -COOH group is also attached to at least two other atoms or groups other than hydrogen; or a compound convertible to said compound during the fermentation process.

Claim 24

A process according to claim 14 wherein R is an alpha-branched C₃-C₈ alkyl, alkenyl, alkynyl, alkoxyalkyl or alkylthioalkyl group; a C₅-C₈ cyclo­alkylalkyl group wherein the alkyl group is an alpha-­branched C₂-C₅ alkyl group; a C₃-C₈ cycloalkyl or C₅-C₈ cycloalkenyl group, either of which may optionally be substituted by methylene or one or more C₁-C₄ alkyl groups or halo atoms; or a 3 to 6 membered oxygen or sulfur containing heterocyclic ring which may be saturated, or fully or partially unsaturated and which may optionally be substituted by one or more C₁-C₄ alkyl groups or halo atoms, or a compound convertible to said compound during the fermentation process.

Claim 25

A process according to claim 14 wherein the compound convertible to substrate RCOOH is R-(CH₂) n -Z wherein R is as previously defined; n is 0, 2, 4 or 6; and Z is -CH₂OH, -CHO, -COOR⁵, -CH₂NH₂ or -CONHR⁶ wherein R⁵ is H or (C 1-6 alkyl; R⁶ is hydrogen, (C 1-4 )­alkyl, -CH(COOH)CH₂COOH, -CH(COOH) (CH₂)₂COOH or -CH(COOH) (CH₂)₂SCH₃ or, when the strain of S . avermitilis lacks only branched-chain amino acid transaminase activity, the compound convertible to substrate RCOOH is R-CO-Z.

Claim 26

A process according to claim 15 wherein R is cyclobutyl cyclopentyl cyclohexyl cycloheptyl 2-methylcyclopropyl 3-cyclohexenyl 1-cyclopentenyl 1-cyclohexenyl 3-methylcyclohexyl (cis/trans) 4-methylenecyclohexyl 3-methylcyclobutyl 3-methylenecyclobutyl 3-cyclopentenyl 1-cyclopropylethyl 3-fluorocyclobutyl 4,4-difluorocyclohexyl isopropyl sec-butyl 2-pentyl 2,3-dimethylpropyl 2-hexyl 2-pent-4-enyl 2-methylthioethyl S-2-methylpentyl R-2-methylpentyl 2-thienyl 3-thienyl 4-tetrahydropyranyl 3-furyl 2-chlorothienyl 3-tetrahydrothienyl 4-methylthio-2-butyl 4-tetrahydrothiopyranyl 4-methoxy-2-butyl or 4-methylthio-2-butyl.

Claim 27

A process according to claim 17 wherein R is cyclopentyl, cyclohexyl or thienyl.

Claim 28

A process according to claim 15 wherein when R is an alpha-branched C₃-C₈ alkyl group, it is not isopropyl or (S)-sec-butyl.

Claim 29

The process according to claim 15 wherein the strain of S . avermitilis ATCC 53567, ATCC 53568, ATCC 53669 or ATCC 53670.

Claim 30

A novel strain belonging to the genus Streptomyces which produces substantially no C-076 when fermented in an aqueous nutrient medium comprising an assimilable source of carbon, nitrogen and inorganic salts under aerobic conditions, said medium being substantially free of an acid of the formula R-COOH , or a compound convertible to said acid by S . avermitilis , wherein R is isopropyl or (S)-sec-butyl, but being capable of producing C-076 upon fermentation in said medium when said medium contains an acid of the formula RCOOH, or a compound convertible to said acid by S . avermitilis , wherein R is isopropyl or (S)-sec-­butyl, wherein said strain is obtained by a process which comprises mutating Streptomyces avermitilis ATCC 31271, ATCC 31272 or mutants thereof.

Claim 31

A novel strain belonging to the genus Streptomyces which produces substantially no C-076 when fermented in an aqueous nutrient medium comprising an assimilable source of carbon, nitrogen and inorganic salts under aerobic conditions, said medium being substantially free of an acid of the formula R-COOH , or a compound convertible to said acid by S . avermitilis , wherein R is isopropyl or (S)-sec-butyl, but being capable of producing C-076 upon fermentation in said medium when said medium contains an acid of the formula RCOOH, or a compound convertible to said acid by S . avermitilis , wherein R is isopropyl or (S)-sec-­butyl, wherein said strain is obtained by a process which comprises mutating Streptomyces avermitilis ATCC 31271, ATCC 31272 or mutants thereof.