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Open Access Research article

Molecular cloning and functional characterization of an ATP-binding cassette transporter OtrC from Streptomyces rimosus

Lan Yu12, Xiangyun Yan1, Long Wang1, Ju Chu1, Yingping Zhuang1, Siliang Zhang1* and Meijin Guo1*

Author Affiliations

1 State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, 130 Meilong Road, Shanghai, 200237, P.R. China

2 School of Life & Science, Huzhou Teachers College, Zhejiang, Huzhou, 313000, P.R. China

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BMC Biotechnology 2012, 12:52  doi:10.1186/1472-6750-12-52

Published: 20 August 2012

Abstract

Background

The otrC gene of Streptomyces rimosus was previously annotated as an oxytetracycline (OTC) resistance protein. However, the amino acid sequence analysis of OtrC shows that it is a putative ATP-binding cassette (ABC) transporter with multidrug resistance function. To our knowledge, none of the ABC transporters in S. rimosus have yet been characterized. In this study, we aimed to characterize the multidrug exporter function of OtrC and evaluate its relevancy to OTC production.

Results

In order to investigate OtrC’s function, otrC is cloned and expressed in E. coli The exporter function of OtrC was identified by ATPase activity determination and ethidium bromide efflux assays. Also, the susceptibilities of OtrC-overexpressing cells to several structurally unrelated drugs were compared with those of OtrC-non-expressing cells by minimal inhibitory concentration (MIC) assays, indicating that OtrC functions as a drug exporter with a broad range of drug specificities. The OTC production was enhanced by 1.6-fold in M4018 (P = 0.000877) and 1.4-fold in SR16 (P = 0.00973) duplication mutants, while it decreased to 80% in disruption mutants (P = 0.0182 and 0.0124 in M4018 and SR16, respectively).

Conclusions

The results suggest that OtrC is an ABC transporter with multidrug resistance function, and plays an important role in self-protection by drug efflux mechanisms. This is the first report of such a protein in S. rimosus, and otrC could be a valuable target for genetic manipulation to improve the production of industrial antibiotics.

Keywords:
Streptomyces rimosus; OtrC; ATP-binding cassette transporter; ATP hydrolysis; Multidrug resistance