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Identification and Characterization of hmr19 Gene Encoding a Multidrug Resistance Efflux Protein from Streptomyces hygroscopicus *** Strain 10-22

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Acta Biochimica et Biophysica Sinica 2004年第8期36卷 519-528页

作者： lei qin , heng-An wang1 , Zhong-qin WU, Xiao-Feng ZHANG, Mei-lei JIN,zi-xin deng2, and guo-ping zhaoshanghaiResearch 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 shanghaiResearch Center of biotechnology Shanghai Institute for Biological Sciences Chinese Academy of Sciences Graduate School of the Chinese Academy of Sciences school of agriculture and biology Shanghai Jiaotong University

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 α-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.

关键词： multidrug resistance (MDR) gene replacement (knockout) Streptomyces Streptomyces hygroscopicus subsp. yingchengensis strain 10-22

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cvhA Gene of Streptomyces hygroscopicus 10-22 Encodes a Negative Regulator for Mycelia Development

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Acta Biochimica et Biophysica Sinica 2006年第4期38卷 271-280页

作者： heng-An wang~(1,2#) lei qin~(2#) ping lu~(1*) zhi-xuan pang~2 zi-xin deng~(3*) guo-ping zhao~(2*) 1 biotechnology institute,school of agriculture and biology,shanghai jiaotong university,shanghai 201101,china 2 Laboratory of Molecular Microbiology,institute of Plant Physiology and Ecology,shanghai institute for Biological Sciences,Chinese Academy of Sciences,shanghai 200032,china 3 Bio-X Life Science Research Center,shanghai jiaotong university,shanghai 200030,china. biotechnologyinstitute SchoolofAgricultureandBiologyShanghaiJiaotongUniversityShanghai201101China LaboratoryofMolecularMicrobiology InstituteofPlantPhysiologyandEcologyShanghaiInstituteforBiologicalSciencesChinese

A five-gene cluster cvhABCDE was identified from Streptomyces hygroscopicus *** thefirst gene of this cluster,cvhA encoded a putative sensor histidine kinase with a predicted sensor domainconsisting of two trans-membrane segments at the N-terminus and a conserved HATPasecdomain at the *** C-terminus polypeptide of CvhA expressed in Escherichia coli was purified and shown to beautophosphorylated with [γ-32P]ATP in *** phosphoryl group was acid-labile and basic-stable,whichsupported histidine as the phosphorylation *** obvious difference of mycelia development was ob-served between the null mutant of cvhA generated by targeted gene replacement and the wild-type parentalstrain 10-22 grown on solid soya flour medium with 2%-8% glucose or sucrose,but the cvhA mutant couldform much more abundant aerial mycelia and spores than the wild-type strain on solid soya flour mediumsupplemented with 6%-8% mannitol,6%-8% sorbitol,4%-6% mannose,or 4%-6% *** pheno-type was complemented by the cloned wild-type cvhA gene,and no difference was observed for growthcurves of the cvhA mutant and the wild strain in liquid minimal medium with the tested sugars at a concen-tration of 4%,6% and 8%.We thus propose that CvhA is likely a sensor histidine kinase and negativelyregulates the morphological differentiation in a sugar-dependent manner in *** 10-22.

关键词： sensor histidine kinase aerial mycelia sugar metabolism

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