Repositioning of a domain in a modular polyketide synthase to promote specific chain cleavage.

Macrocyclic polyketides exhibit an impressive range of medically useful activities, and there is great interest in manipulating the genes that govern their synthesis. The 6-deoxyerythronolide B synthase (DEBS) of Saccharopolyspora erythraea, which synthesizes the aglycone core of the antibiotic erythromycin A, has been modified by repositioning of a chain-terminating cyclase domain to the carboxyl-terminus of DEBS1, the multienzyme that catalyzes the first two rounds of polyketide chain extension. The resulting mutant markedly accelerates formation of the predicted triketide lactone, compared to a control in which the repositioned domain is inactive. Repositioning of the cyclase should be generally useful for redirecting polyketide synthesis to obtain polyketides of specified chain lengths.

[1]  D. Sherman,et al.  Molecular genetics of polyketides and its comparison to fatty acid biosynthesis. , 1990, Annual review of genetics.

[2]  M. Staver,et al.  Biosynthesis of the erythromycin macrolactone and a rational approach for producing hybrid macrolides. , 1992, Gene.

[3]  D. Hopwood,et al.  Nucleotide sequence and deduced functions of a set of cotranscribed genes of Streptomyces coelicolor A3(2) including the polyketide synthase for the antibiotic actinorhodin. , 1992, The Journal of biological chemistry.

[4]  A. Joshi,et al.  Construction, expression, and characterization of a mutated animal fatty acid synthase deficient in the dehydrase function. , 1993, The Journal of biological chemistry.

[5]  R. Potenz,et al.  Organization of a cluster of erythromycin genes in Saccharopolyspora erythraea , 1990, Journal of bacteriology.

[6]  P. Leadlay,et al.  An unusually large multifunctional polypeptide in the erythromycin-producing polyketide synthase of Saccharopolyspora erythraea , 1990, Nature.

[7]  P. Leadlay,et al.  Limited proteolysis and active-site studies of the first multienzyme component of the erythromycin-producing polyketide synthase. , 1994, Journal of Biological Chemistry.

[8]  J B McAlpine,et al.  An erythromycin analog produced by reprogramming of polyketide synthesis. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[9]  J. Staunton.  THE EXTRAORDINARY ENZYMES INVOLVED IN ERYTHROMYCIN BIOSYNTHESIS , 1991 .

[10]  C. Kao,et al.  ENGINEERED BIOSYNTHESIS OF A TRIKETIDE LACTONE FROM AN INCOMPLETE MODULAR POLYKETIDE SYNTHASE , 1994 .

[11]  P. Leadlay,et al.  Stereospecific acyl transfers on the erythromycin-producing polyketide synthase. , 1994, Science.

[12]  P. Leadlay,et al.  An acyl-carrier-protein-thioesterase domain from the 6-deoxyerythronolide B synthase of Saccharopolyspora erythraea. High-level production, purification and characterisation in Escherichia coli. , 1991, European journal of biochemistry.

[13]  J B McAlpine,et al.  Modular organization of genes required for complex polyketide biosynthesis. , 1991, Science.

[14]  P. Leadlay,et al.  Heterologous expression in Escherichia coli of an intact multienzyme component of the erythromycin-producing polyketide synthase. , 1993, European journal of biochemistry.

[15]  J. Butte,et al.  Actinomycete antibiotics. III. The biogenesis of erythronolide, the C-21 branched chain lactone in erythromycin. , 1962, The Journal of biological chemistry.

[16]  J. Lawrence,et al.  Isolation and sequence analysis of polyketide synthase genes from the daunomycin-producing Streptomyces sp. strain C5 , 1994, Journal of bacteriology.

[17]  P. Leadlay,et al.  Identification of DEBS 1, DEBS 2 and DEBS 3, the multienzyme polypeptides of the erythromycin‐producing polyketide synthase from Saccharopolyspora erythraea , 1992, FEBS letters.

[18]  S. Ripka,et al.  The multifunctional 6-methylsalicylic acid synthase gene of Penicillium patulum. Its gene structure relative to that of other polyketide synthases. , 1990, European journal of biochemistry.