This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Kim, E. S. Articles by Sherman, D. H. Search for Related Content PubMed PubMed Citation Articles by Kim, E. S. Articles by Sherman, D. H.

 Previous Article  |  Next Article 

J. Bacteriol., Mar 1995, 1202-1207, Vol 177, No. 5
Copyright © 1995, American Society for Microbiology

Heterologous expression of an engineered biosynthetic pathway: functional dissection of type II polyketide synthase components in Streptomyces species

ES Kim, KD Cramer, AL Shreve and DH Sherman
Department of Microbiology, University of Minnesota, St. Paul 55108.

Polyketides are an extensive class of secondary metabolites with diverse molecular structures and biological activities. A plasmid-based multicomponent polyketide synthase expression cassette was constructed using a subset of actinorhodin (act) biosynthetic genes (actI-orf1, actI-orf2, actI-orf3, actIII, actVII, and actIV) from Streptomyces coelicolor which specify the construction of the anthraquinone product aloesaponarin II, a molecule derived from acetyl coenzyme A and 7 malonyl coenzyme A extender units. This system was designed as an indicator pathway in Streptomyces parvulus to quantify polyketide product formation and to examine the functional significance of specific polyketide synthase components, including the act beta- ketoacyl synthase (beta-KS; encoded by actI-orf1 and actI-orf2) and the act cyclase/dehydrase (encoded by actVII-orf4). Site-directed mutagenesis of the putative active site Cys (to a Gln) in the actI-orf1 beta-KS product completely abrogated aloesaponarin II production. Changing the putative acyltransferase active-site Ser (to a Leu) located in the actI-orf1 beta-KS product led to significantly reduced but continued production of aloesaponarin II. Replacement of the expression cassette with one containing a mutant form of actI-orf2 gave no production of aloesaponarin II or any other detectable polyketide products. However, an expression cassette containing a mutant form of actVII-orf4 gave primarily mutactin with low-level production of aloesaponarin II.

This article has been cited by other articles:

• Fang, J., Zhang, Y., Huang, L., Jia, X., Zhang, Q., Zhang, X., Tang, G., Liu, W. (2008). Cloning and Characterization of the Tetrocarcin A Gene Cluster from Micromonospora chalcea NRRL 11289 Reveals a Highly Conserved Strategy for Tetronate Biosynthesis in Spirotetronate Antibiotics. J. Bacteriol. 190: 6014-6025 [Abstract] [Full Text]  
• Kim, D.-J., Huh, J.-H., Yang, Y.-Y., Kang, C.-M., Lee, I.-H., Hyun, C.-G., Hong, S.-K., Suh, J.-W. (2003). Accumulation of S-Adenosyl-L-Methionine Enhances Production of Actinorhodin but Inhibits Sporulation in Streptomyces lividans TK23. J. Bacteriol. 185: 592-600 [Abstract] [Full Text]  
• Pfeifer, B. A., Khosla, C. (2001). Biosynthesis of Polyketides in Heterologous Hosts. Microbiol. Mol. Biol. Rev. 65: 106-118 [Abstract] [Full Text]  
• He, M., Varoglu, M., Sherman, D. H. (2000). Structural Modeling and Site-Directed Mutagenesis of the Actinorhodin beta -Ketoacyl-Acyl Carrier Protein Synthase. J. Bacteriol. 182: 2619-2623 [Abstract] [Full Text]  
• Jez, J. M., Noel, J. P. (2000). Mechanism of Chalcone Synthase. pKa OF THE CATALYTIC CYSTEINE AND THE ROLE OF THE CONSERVED HISTIDINE IN A PLANT POLYKETIDE SYNTHASE. J. Biol. Chem. 275: 39640-39646 [Abstract] [Full Text]