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ISSN
1672-9145
Acta Biochim Biophys Sin 2004, 36(8): 519–528
CN 31-1940/Q
Identification and Characterization of hmr19 Gene Encoding a
Multidrug Resistance Efflux Protein from Streptomyces hygroscopicus subsp.
yingchengensis Strain 10-22
Lei QIN#, Heng-An WANG1#, Zhong-Qin WU, Xiao-Feng ZHANG, Mei-Lei JIN, Zi-Xin DENG2,
and Guo-Ping ZHAO*
Shanghai Research Center of Biotechnology, Shanghai Institute
for Biological Sciences, Chinese Academy of Sciences, Graduate School of the
Chinese Academy of Sciences, Shanghai 200233, China; 1School of Agriculture and Biology, Shanghai
Jiaotong University, Shanghai 201101, China; 2School of Life Science and Biotechnology,
Shanghai Jiaotong University, Shanghai 200030, China
Abstract The hmr19 gene
was cloned from Streptomyces hygroscopicus subsp. yingchengensis
strain 10-22, a bacterium strain producing agricultural antibiotics. Sequence
similarity comparison indicates that hmr19 gene may encode a predicted
protein with 14 putative transmembrane a-helical
spanners, belonging to the drug:H+ antiporter-2 family of the major
facilitator superfamily. The expression of hmr19 in the mycelium
of strain 10-22 was detected by Western blotting analysis. Gene replacement
technology was employed to construct an hmr19 disruption mutant. The
growth inhibition test against different antibiotics indicated that the mutant
strain was 5–20
fold more susceptible to tetracycline, vancomycin and mitomycin C than the
parental wild type strain. The mutant took up tetracycline much faster and
accumulated more antibiotics than the wild type strain 10-22. While with the
addition of an energy uncoupler, carbonyl cyanide m-chlorophenylhydrazone,
the characteristics of the accumulation of [3H]tetracycline in these two
strains were almost the same. It was thus concluded that hmr19 encoded a
multidrug resistance efflux protein.
Key words multidrug resistance
(MDR); gene replacement (knockout); Streptomyces; Streptomyces
hygroscopicus subsp. yingchengensis strain 10-22
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Received: March 18, 2004 Accepted: June 18, 2004
This work was supported by the grants from the National Natural Science
Foundation of China (No. 30070016) and the National High Technology Research
and Development Program of China (No. 2001AA234051)
#These authors contributed equally to this
work
*Corresponding author: Tel, 86-21-54971046; Fax, 86-21-64837495;
E-mail, [email protected]