Shanghai Laiyi Center for Biopharmaceutical R and D

Shanghai, China

Shanghai Laiyi Center for Biopharmaceutical R and D

Shanghai, China
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Liu J.-J.,Shanghai JiaoTong University | Rao M.,Shanghai Laiyi Center for Biopharmaceutical R and D | Ge M.,Shanghai Laiyi Center for Biopharmaceutical R and D | Wei W.,Shanghai JiaoTong University | Qian X.-P.,Shanghai JiaoTong University
Chinese Journal of Antibiotics | Year: 2014

Objective: To preliminarily identify a new lipopeptide antibiotics, optimize its fermentation conditions and detect its bioactivity. Methods: Preliminarily identify the new lipopeptide antibiotics by ultra-performance liquid chromatography coupled with tandem quadrupole and time of flight high-resolution mass spectrometry (UPLC-Q-TOF-HRMS). Add different precursor substances to the fermentation medium, detect the target product yield by HPLC method and ultimately determine the optimal fermentation conditions. Detect the antimicrobial activity using the disc diffusion method. Result: Preliminary results indicate that the new lipopeptide compound is a new arylomycin A6. The highest yield was obtained when 3g/L isoleucine was applied. Conclusion: arylomycin A2, A4 and compound C exhibit antibacterial activities against Sarcina lutea and Staphylococcus epidermidis, and compound C showed the strongest antibacterial activity amongst them.


Wan G.-Q.,China Pharmaceutical University | Wan G.-Q.,Shanghai Laiyi Center for Biopharmaceutical R and D | Yang Z.-J.,Shanghai Laiyi Center for Biopharmaceutical R and D | Yang Z.-J.,Shanghai JiaoTong University | And 5 more authors.
Chinese Journal of Antibiotics | Year: 2014

Objective: To discover polyketide and macrolide compounds from metabolites of actinomycetes. Methods: Type I polyketidesynthase genes was cloned from genomes of actinomycetes by PCR technology, then metabolites of actinomycetes were analyzed by UPLC-MS/MS method. Results: 18 positive strains were screened from 40 strains by PCR technology. Seven macrotetrolides were discovered from the strain HCCB11494 by UPLCMS/MS analysis. And this strain was preliminarily identified as Streptomyces parvus according to gyrB sequences. Conclusion: Macrolide compounds could be discovered from actinomycetes by PCR technology in combination with UPLC-MS analysis.


Jin X.,Nanjing Agricultural University | Jin X.,Shanghai Laiyi Center for Biopharmaceutical R and D | Wei W.,Shanghai Laiyi Center for Biopharmaceutical R and D | Wei W.,Shanghai JiaoTong University | And 6 more authors.
Chinese Journal of Antibiotics | Year: 2014

Objective To investigate the influence of A21978C biosynthesis gene clusters of 3 regulatory genes dptR1, dptR2 and dptR3 on synthesis of A1978C. Methods dptR1, dptR2 and dptR3 were deleted by homologous recombination and the production of A21978C in the corresponding mutants was detected via HPLC. Results There were three ΔdptR1 mutants, four ΔdptR2 mutants and two ΔdptR3 mutants obtained. The production of A21978C in ΔdptR1 and ΔdptR2 mutants was at a similar level to that of the wild-type strain, while the ΔdptR3 mutant lost its ability to produce A21978C. Conclusion dptR3 was essential for A21978C production and was a positive regulatory gene.


Yang S.,East China University of Science and Technology | He H.,Institutes of Plant Physiology and Ecology | Li Z.,Shanghai Laiyi Center for Biopharmaceutical R and D | Shen Y.,Institutes of Plant Physiology and Ecology | And 2 more authors.
Chinese Journal of Antibiotics | Year: 2013

ECO-0501 is a new linear polyene found from Amycolatopsis orientalis. Because of the strong bacteridicial activity of ECO-0501 and its low toxity, preclinical development has been started on this antibiotic. Four positive cosmid clones were identified by colony hybridization that cover the whole ECO-0501 locus, including NL5B, NL9-3-1, NL3-1C and NL3-3A. Sequence analysis of the gene ORF4 revealed that its deduced protein might be a positive regulator controlling ECO-0501 biosynthesis. Overexpression of ORF4 showed that the yield of ECO-0501 increased about six fold than that of wild-type strain. The present results provide us a tool for further study of ECO-0501 development and its regulatory mechanism.


Li Y.-M.,China Pharmaceutical University | Li Y.-M.,Shanghai Laiyi Center for Biopharmaceutical R and D | Xu J.-Y.,China Pharmaceutical University | Xu L.,Shanghai Laiyi Center for Biopharmaceutical R and D | And 4 more authors.
Chinese Journal of Pharmaceutical Biotechnology | Year: 2013

The objective of this study was heterologous expression of the glycosyltranferase gene bgtfA from balhimycin producing-strain in Amycolatopsis orientalis, a vancomycin producing strain, to obtain hybride vancomycin from the fermentation broth of the constructed engineered strain. To obtain an engineered strain, the gene of bgtfA was coloned into vector pULVK2AE, and then the recombinant vector pLYEBA2AE was transformed into Amycolatopsis orientalis by electroporation. The broth of engineered strain was detected and analyzed by HPLC and LC-MS. The results showed that heterologous bgtfA gene was successfully expressed in Amycolatopsis orientalis with the replicative plasmid pLYEBA2AE by electroporation and an expected hybride vancomycin named LYV07ww01 was detected and identified by HPLC and LC-MS in the broth of engineered mutant. This indicated that BgtfA has its broad substrate specificity not only to transfer epivancosamine to balhimycin aglycone but also to transfer vancosamine to vancomycin aglycone.


Pang L.,East China University of Science and Technology | Pang L.,Shanghai Laiyi Center for Biopharmaceutical R and D | Wei W.,Shanghai Laiyi Center for Biopharmaceutical R and D | Wei W.,Shanghai JiaoTong University | And 5 more authors.
Chinese Journal of Antibiotics | Year: 2015

Objective To discover some relationships between tfpA and the biosynthesis of A21978C which belongs to the TetR-family, a transcriptional regulator. Methods tfpA was deleted by homologous recombination, and the production of A21978C in the mutants was detected via HPLC. Then complementation and over-expression were used to confirm the function of this gene. Results Compared to the original strain, production of A21978C in the ΔtfpA mutant increased obviously. While the O-tfpA mutant lost its ability to produce A21978C, and C-tfpA mutant was at a similar level to that of the original strain. Conclusion tfpA exerts a strong influence on A21978C production and is a negative regulatory gene.


Zhu L.,Shanghai JiaoTong University | Zhu L.,Shanghai Laiyi Center for Biopharmaceutical R and D | Rao M.,Shanghai Laiyi Center for Biopharmaceutical R and D | Luo M.-Y.,Shanghai Laiyi Center for Biopharmaceutical R and D
Chinese Journal of Antibiotics | Year: 2015

Objective To introduce the I-SceI restriction modification system into the Amycolatopsis orientalis for high efficiency recombination by DNA double strand break with I-SceI. Methods Firstly, the shuttle vector pLYZL102 with 18bp I-SceI recognized sites and the plasmid pLYZL104 with sceS gene following ermP promoter were constructed, and the up and down stream fragments of hemA2 gene for ECO-0501 synthesis were inserted into pLYZL102. The resulted plasmid pLYhemA2D was electrotransported to A. orientalis and the single cross-over transformants were obtained. Then pLYZL104 was introduced into the single cross-over strain and the efficiency of the homologous recombination was analyzed before and after the sceS expression. Subsequently, the fermentation products of ΔhemA2 mutants were assayed by HPLC. Results Expression of sceS gene in A. orientalis resulted in promotion of homologous recombination and markerless deletion of hemA2. Conclusion I-SceI toolkit was useful for markless genetic manipulation in A. orientalis, and hemA2 was one of key genes for ECO-0501 synthesis.


Zhang Y.,Shanghai JiaoTong University | Niu Y.,Shanghai JiaoTong University | Luo Y.,University of Maryland University College | Ge M.,Shanghai Laiyi Center for Biopharmaceutical R and D | And 4 more authors.
Food Chemistry | Year: 2014

Thymol-loaded zein nanoparticles stabilized with sodium caseinate (SC) and chitosan hydrochloride (CHC) were prepared and characterized. The SC stabilized nanoparticles had well-defined size range and negatively charged surface. Due to the presence of SC, the stabilized zein nanoparticles showed a shift of isoelectric point from 6.18 to 5.05, and had a desirable redispersibility in water at neutral pH after lyophilization. Coating with CHC onto the SC stabilized zein nanoparticles resulted in increased particle size, reversal of zeta potential value from negative to positive, and improved encapsulation efficiency. Both thymol-loaded zein nanoparticles and SC stabilized zein nanoparticles had a spherical shape and smooth surface, while the surfaces of CHC-SC stabilized zein nanoparticles seemed rough and had some clumps. Encapsulated thymol was more effective in suppressing gram-positive bacterium than un-encapsulated thymol for a longer time period. © 2013 Elsevier Ltd. All rights reserved.


Yang Z.,Shanghai Institute of Pharmaceutical Industry | Ge M.,Shanghai Laiyi Center for Biopharmaceutical R and D | Yin Y.,Shanghai Laiyi Center for Biopharmaceutical R and D | Chen Y.,Shanghai Normal University | And 2 more authors.
Chemistry and Biodiversity | Year: 2012

A novel phytotoxic nonenolide, (6S,7R,9R)-6,7-dihydroxy-9-propylnon-4-eno- 9-lactone (1), was isolated from solid cultures of the endophytic fungus Phomopsis sp. HCCB03520, together with three known compounds, cytochalasin H (2), cytochalasin N (3), and epoxycytochalasin H (4). The structures of these compounds were elucidated through spectroscopic analysis, and the absolute configurations were determined by CD spectroscopy. Phytotoxic activities of compounds 1-4 were also investigated. Compound 1 showed phytotoxic activity on germination and radicle growth of Medicago sativa, Trifolium hybridum, and Buchloe dactyloides. © 2012 Verlag Helvetica Chimica Acta AG, Zürich.


Xu L.,Nanjing Agricultural University | Xu L.,Shanghai Laiyi Center for Biopharmaceutical R and D | Huang H.,CAS Institute of Plant Physiology and Ecology | Wei W.,Shanghai Laiyi Center for Biopharmaceutical R and D | And 20 more authors.
BMC Genomics | Year: 2014

Background: Amycolatopsis orientalis is the type species of the genus and its industrial strain HCCB10007, derived from ATCC 43491, has been used for large-scale production of the vital antibiotic vancomycin. However, to date, neither the complete genomic sequence of this species nor a systemic characterization of the vancomycin biosynthesis cluster (vcm) has been reported. With only the whole genome sequence of Amycolatopsis mediterranei available, additional complete genomes of other species may facilitate intra-generic comparative analysis of the genus.Results: The complete genome of A. orientalis HCCB10007 comprises an 8,948,591-bp circular chromosome and a 33,499-bp dissociated plasmid. In total, 8,121 protein-coding sequences were predicted, and the species-specific genomic features of A. orientalis were analyzed in comparison with that of A. mediterranei. The common characteristics of Amycolatopsis genomes were revealed via intra- and inter-generic comparative genomic analyses within the domain of actinomycetes, and led directly to the development of sequence-based Amycolatopsis molecular chemotaxonomic characteristics (MCCs). The chromosomal core/quasi-core and non-core configurations of the A. orientalis and the A. mediterranei genome were analyzed reciprocally, with respect to further understanding both the discriminable criteria and the evolutionary implementation. In addition, 26 gene clusters related to secondary metabolism, including the 64-kb vcm cluster, were identified in the genome. Employing a customized PCR-targeting-based mutagenesis system along with the biochemical identification of vancomycin variants produced by the mutants, we were able to experimentally characterize a halogenase, a methyltransferase and two glycosyltransferases encoded in the vcm cluster. The broad substrate spectra characteristics of these modification enzymes were inferred.Conclusions: This study not only extended the genetic knowledge of the genus Amycolatopsis and the biochemical knowledge of vcm-related post-assembly tailoring enzymes, but also developed methodology useful for in vivo studies in A. orientalis, which has been widely considered as a barrier in this field. © 2014 Xu et al.

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