Applied Microbiology and Biotechnology

, Volume 89, Issue 6, pp 1709–1719

Titer improvement of iso-migrastatin in selected heterologous Streptomyces hosts and related analysis of mRNA expression by quantitative RT–PCR

Biotechnological Products and Process Engineering

DOI: 10.1007/s00253-010-3025-1

Cite this article as:
Yang, D., Zhu, X., Wu, X. et al. Appl Microbiol Biotechnol (2011) 89: 1709. doi:10.1007/s00253-010-3025-1

Abstract

iso-Migrastatin (iso-MGS) has been actively pursued recently as an outstanding candidate of antimetastasis agents. Having characterized the iso-MGS biosynthetic gene cluster from its native producer Streptomyces platensis NRRL 18993, we have recently succeeded in producing iso-MGS in five selected heterologous Streptomyces hosts, albeit the low titers failed to meet expectations and cast doubt on the utility of this novel technique for large-scale production. To further explore and capitalize on the production capacity of these hosts, a thorough investigation of these five engineered strains with three fermentation media for iso-MGS production was undertaken. Streptomyces albus J1074 and Streptomyces lividans K4-114 were found to be preferred heterologous hosts, and subsequent analysis of carbon and nitrogen sources revealed that sucrose and yeast extract were ideal for iso-MGS production. After the initial optimization, the titers of iso-MGS in all five hosts were considerably improved by 3–18-fold in the optimized R2YE medium. Furthermore, the iso-MGS titer of S. albus J1074 (pBS11001) was significantly improved to 186.7 mg/L by a hybrid medium strategy. Addition of NaHCO3 to the latter finally afforded an optimized iso-MGS titer of 213.8 mg/L, about 5-fold higher than the originally reported system. With S. albus J1074 (pBS11001) as a model host, the expression of iso-MGS gene cluster in four different media was systematically studied via the quantitative RT–PCR technology. The resultant comparison revealed the correlation of gene expression and iso-MGS production for the first time; synchronous expression of the whole gene cluster was crucial for optimal iso-MGS production. These results reveal new insights into the iso-MGS biosynthetic machinery in heterologous hosts and provide the primary data to realize large-scale production of iso-MGS for further preclinical studies.

Keywords

iso-Migrastatin Heterologous Streptomyces hosts Fermentation optimization Quantitative RT–PCR 

Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  1. 1.Department of Chemical and Biological Engineering, Institute of BioengineeringZhejiang UniversityHangzhouChina
  2. 2.Division of Pharmaceutical Science, School of PharmacyUniversity of Wisconsin–MadisonMadisonUSA