Geosmin Biosynthesis in Streptomyces avermitilis. Molecular Cloning, Expression, and Mechanistic Study of the Germacradienol/Geosmin Synthase

Geosmin ( 1 ) is responsible for the characteristic odor of moist soil. The Gram-positive soil bacterium Streptomyces avermitilis produces geosmin ( 1 ) as well as its precursor germacradienol ( 3 ). The S. avermitilis gene SAV2163 ( geoA ) is extremely similar to the S. coelicolor A3(2) SCO6073 gene that encodes a germacradienol/geosmin synthase. S. avermitilis mutants with a deleted geoA were unable to produce either germacradienol ( 3 ) or geosmin ( 1 ). Biosynthesis of both compounds was restored by introducing an intact geoA gene into the mutants. Incubation of recombinant GeoA, encoded by the SAV2163 gene of S. avermitilis , with farnesyl diphosphate ( 2 ) in the presence of Mg 2+ gave a mixture of (4 S ,7 R )-germacra-1(10) E ,5 E -diene-11-ol ( 3 ) (66%), (7 S )-germacrene D ( 4 ) (24%), geosmin ( 1 ) (8%), and a hydrocarbon, tentatively assigned the structure of octalin 5 (2%). Incubation of this germacradienol/geosmin synthase with [1,1- 2 H 2 ]FPP ( 2a ) gave geosmin-d 1 ( 1a ), as predicted. When recombinant GeoA from either S. avermitilis or S. coelicolor A3(2) was incubated with nerolidyl diphosphate ( 8 ), only the acyclic elimination products β-farnesene ( 10 ), ( Z )-α-farnesene ( 11 ), and ( E )-α-farnesene ( 12 ) were formed, thereby ruling out nerolidyl diphosphate as an intermediate in the conversion of farnesyl diphosphate to geosmin, germacradienol, and germacrene D.