| Reference | [1]. J Appl Microbiol. 2015 Oct;119(4):1064-74. doi: 10.1111/jam.12919.<br />
Co-overexpression of lmbW and metK led to increased lincomycin A production and decreased byproduct lincomycin B content in an industrial strain of Streptomyces lincolnensis.<br />
Pang AP(1)(2)(3), Du L(1)(2)(3), Lin CY(1)(2)(3), Qiao J(1)(2)(3), Zhao GR(1)(2)(3).<br />
Author information: (1)Key Laboratory of Systems Bioengineering, Ministry of Education, Tianjin, China. (2)Department of Pharmaceutical Engineering, School of Chemical Engineering and Technology, Tianjin University, Tianjin, China. (3)SynBio Research Platform, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin, China.<br />
AIMS: To improve lincomycin A production and decrease the content of byproduct lincomycin B in an industrial lincomycin-producing strain. METHODS AND RESULTS: The in silico analysis indicated that LmbW could be involved in propylproline biosynthesis of lincomyin A. In this study, we constructed an lmbW deletion mutant and found that the mutant lost the ability to produce lincomycin A, but increased the accumulation of lincomycin B. The loss of lincomycin A production can be restored by complementing the mutant with the expression of lmbW gene. When lmbW and metK (encoding S-adenosylmethionine synthetase) was co-overexpressed, lincomycin A titre was 1744·6 mg l(-1) , a 35·83% improvement over the original strain. Meanwhile, the content of lincomycin B was reduced to 4·41%, a remarkable decrease of 34·76%, compared to that of the original strain. CONCLUSIONS: lmbW encodes a C-methyltransferase involved in the biosynthesis of lincomycin A but not lincomycin B. Co-overexpression of lmbW and metK improved lincomycin A production and decreased the content of lincomycin B. SIGNIFICANCE AND IMPACT OF THE STUDY: The engineered Streptomyces lincolnensis strain shows promising application in the fermentation production of lincomycin A, which may help cut production costs and simplify downstream separation processes.<br />
DOI: 10.1111/jam.12919 PMID: 26248490 [Indexed for MEDLINE]<br />
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[2]. J Assoc Off Anal Chem. 1984 May-Jun;67(3):582-8.<br />
Gas chromatographic-mass spectrometric detection and quantitation of lincomycin in animal feedingstuffs.<br />
McMurray CH, Blanchflower WJ, Rice DA.<br />
A substantially improved assay was developed for lincomycin A in animal feedingstuffs. The assay allows unambiguous quantitation of at least 0.1 ppm in feed. Lincomycin B did not interfere because of differences in both retention time and mass of the main fragment ion in electron impact (EI) spectra. The assay using single ion monitoring with EI detection would not discriminate between lincomycin A and clindamycin. The presence of the latter was easily confirmed by using gas chromatography-mass spectrometry in the chemical ionization mode. The assay for lincomycin A was linear in the range 0-40 ng applied to the gas chromatographic column. The recovery was 93.4 +/- 4.2% at 1 and 5 ppm and 86.2 +/- 5.5% at 0.1 ppm in feed. The coefficient of variation of the assay was 4.8% at both 1 and 5 ppm, and was 6.43% at 0.1 ppm.<br />
PMID: 6547713 [Indexed for MEDLINE]<br />
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[3]. J Pharm Biomed Anal. 1999 Sep;20(5):745-52. doi: 10.1016/s0731-7085(99)00043-6.<br />
Liquid chromatography method for separation of clindamycin from related substances.<br />
Orwa JA(1), Vandenbempt K, Depuydt S, Roets E, Hoogmartens J.<br />
Author information: (1)Katholieke Universiteit Leuven, Faculteit Farmaceutische Wetenschappen, Laboratorium voor Farmaceutische Chemie en Analyse van Geneesmiddelen, Belgium.<br />
A reversed-phase liquid chromatography method has been developed for the separation of clindamycin from 7-epiclindamycin, clindamycin B, lincomycin, lincomycin B, 7-epilincomycin and other impurities of unknown identity. The method uses a Hypersil ODS, 5 microm, 250 x 4.6 mm i.d. column maintained at 45 degrees C. The mobile phase comprises acetonitrile phosphate buffer (1.35% v/v phosphoric acid, adjusted to pH 6.0 with ammonium hydroxide)-water (35:40:25, v/v) at a flow rate of 1.0 ml/min. UV detection is performed at 210 nm. The method was tested on several C-18 columns and showed good robustness. Robustness was further evaluated by performing a full-fraction factorial design experiment. The method showed good selectivity, linearity, and repeatability. It is also suitable for analysis of clindamycin formulations.<br />
DOI: 10.1016/s0731-7085(99)00043-6 PMID: 10701982 [Indexed for MEDLINE]<br />
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[4]. AMB Express. 2020 Aug 20;10(1):152. doi: 10.1186/s13568-020-01089-1.<br />
The inhibitory effects of metabolites from Bacillus pumilus on potato virus Y and the induction of early response genes in Nicotiana tabacum.<br />
Shen S(1)(2)(3)(4)(5), Li W(6)(7)(8)(9).<br />
Author information: (1)Guizhou University, Guiyang, 550025, Guizhou, China. (2)Academy of Agriculture and Forestry Sciences, Qinghai University, 810016, Xining, Qinghai, China. (3)Key Laboratory of Potato Breeding of Qinghai Province, Xining, 810016, Qinghai, China. (4)State Key Laboratory of Plateau Ecology and Agriculture, Xining, 810016, Qinghai, China. (5)The Tibet Plateau Biotechnology Key Lab of Ministry of Education, Xining, 810016, Qinghai, China. (6)Academy of Agriculture and Forestry Sciences, Qinghai University, 810016, Xining, Qinghai, China. [email protected]. (7)Key Laboratory of Potato Breeding of Qinghai Province, Xining, 810016, Qinghai, China. [email protected]. (8)State Key Laboratory of Plateau Ecology and Agriculture, Xining, 810016, Qinghai, China. [email protected]. (9)The Tibet Plateau Biotechnology Key Lab of Ministry of Education, Xining, 810016, Qinghai, China. [email protected].<br />
To develop a new antiviral preparation from a microbial source, the halophilic bacterium Bacillus pumilus E303035 was isolated from a soil sample collected at Qarhan Salt Lake in Qinghai, China. The inhibitory activity of an ethyl acetate extract of its fermentation broth was higher than that of an n-butanol extract. After isolation and purification, 9 compounds were obtained: cyclo(L-Leu-L-Pro) (1), cyclo(L-Pro-L-Tyr) (2), Brevianamide F (3), 2-(3-Indolyl) ethanol (4), N-[2-(1H-indol-3-yl) ethyl] acetamide (5), 3, 3-di(1H-indol-3-yl)propane-1,2-diol (6), Lincomycin B (7), dibutylphthalate (8), and p-hydroxyphenethyl alcohol (9). Compounds 1, 5, and 9 showed inhibitory activities against potato virus Y (PVY). Compounds 1, 4, and 9 had significant inhibitory activity against genes HC-pro, P3, and Nib, compound 5 against gene P3, and compounds 1 and 4 against NIa. Compounds 1, 4, 5, and 9 had significant inhibitory activity against genes VPg and 6K1. Active compounds 1, 5, and 9 had various effects on the expression of viral genes related to pathogenesis. Expression of genes cullin and XTH was up-regulated and CP was down-regulated, compared to the positive control. In conclusion, compounds 1, 5, and 9 might be considered as potential antiviral agents for future development.<br />
DOI: 10.1186/s13568-020-01089-1 PMCID: PMC7441133 PMID: 32816147<br />
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[5]. Prep Biochem Biotechnol. 2020;50(6):529-537. doi: 10.1080/10826068.2019.1710714. Epub 2020 Jan 9.<br />
Amplification of lmbB1 gene in Streptomyces lincolnensis improves quantity and quality of lincomycin A fermentation.<br />
Yang J(1), Ye R(1), Zhang H(2), Liu Y(2).<br />
Author information: (1)The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai, China. (2)Topfond Pharmaceutical Co., Ltd, Henan, China.<br />
As a lincosamide antibiotic, lincomycin is still important for treating diseases caused by Gram-positive bacteria. Manufacturing of lincomycin needs efforts to, e.g. maximize desirable species and minimizing unwanted fermentation byproducts. Analysis of the lincomycin biosynthetic gene cluster of Streptomyces lincolnensis, lmbB1, was shown to catalyze the conversion of L-dopa but not of L-tyrosine and then further generated the precursor of lincomycin A. Based on the principle of directed breeding, a strain termed as S. lincolnensis 24-2, was obtained in this work. By overexpressing the lmbB1 gene, this strain produces efficacious lincomycin A and suppresses melanin generation, whereas contains unwanted lincomycin B. The good fermentation performance of the mutant-lmbB1 (M-lmbB1) was also confirmed in a 15 L-scale bioreactor, which increased the lincomycin A production by 37.6% compared with control of 6435 u/mL and reduced the accumulation of melanin by 29.9% and lincomycin B by 73.4%. This work demonstrated that the amplification of lmbB1 gene mutation and metabolic engineering could promote lincomycin biosynthesis and might be helpful for reducing the production of other industrially unnecessary byproduct.<br />
DOI: 10.1080/10826068.2019.1710714 PMID: 31916478 [Indexed for MEDLINE]
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