Enhancing tylosin production by combinatorial overexpression of efflux,SAM biosynthesis,and regulatory genes in hyperproducing Streptomyces xinghaiensis strain

Tylosin is a 16-membered macrolide antibiotic widely used in veterinary medicine to control infections caused by Gram-positive pathogens and mycoplasmas.To improve the fermentation titer of tylosin in the hyperproducing Streptomyces xinghaiensis strain TL01,we sequenced its whole genome and identified the biosynthetic gene cluster therein.Overexpression of the tylosin efflux gene tlrC,the cluster-situated S-adenosyl methionine(SAM)synthetase gene metK_(cs),the SAM biosynthetic genes adoK_(cs)-metFcs,or the pathway-specific activator gene tylR enhanced tylosin production by 18%,12%,11%,and 11%in the respective engineered strains TLPH08-2,TLPH09,TLPH10,and TLPH12.Co-overexpression of metK_(cs)and adoK_(cs)-metFcs as two transcripts increased tylosin production by 22%in the resultant strain TLPH11 compared to that in TL01.Furthermore,combinational overexpression of tlrC,metK_(cs),adoK_(cs)-metFcs,and tylR as four transcripts increased tylosin production by 23%(10.93g/L)in the resultant strain TLPH17 compared to that in TL01.However,a negligible additive effect was displayed upon combinational overexpression in TLPH17 as suggested by the limited increment of fermentation titer compared to that in TLPH08-2.Transcription analyses indicated that the expression of tlrC and three SAM biosynthetic genes in TLPH17 was considerably lower than that of TLPH08-2 and TLPH11.Based on this observation,the five genes were rearranged into one or two operons to coordinate their overexpression,yielding two engineered strains TLPH23 and TLPH24,and leading to further enhancement of tylosin production over TLPH17.In particular,the production of TLPH23 reached 11.35 g/L.These findings indicated that the combinatorial strategy is a promising approach for enhancing tylosin production in high-yielding industrial strains.

基于Gabor变换和组稀疏表示的敦煌壁画修复算法

在敦煌壁画修复过程中,初始字典的随机选取易陷入局部最优,仅以颜色欧氏距离作为图像块分组标准会导致图像修复后易出现结构模糊和线条不连续等问题。针对以上问题,提出了一种基于Gabor变换和组稀疏表示的敦煌壁画修复算法。首先,采用互信息作为图像块分组准则,并建立相似结构组,这使得组稀疏表示更加合理;然后,通过Gabor小波变换对相似结构组进行特征信息提取,并结合PCA降维的方式得到初始化结构组的特征字典,避免了字典初始化随机选取的不足;最后,采用奇异值SVD分解和分裂Bregman迭代优化方法对结构组字典和稀疏系数进行学习并完成壁画图像的修复。实验结果表明,相比于其他对比算法,所提方法取得了较好的主客观修复效果。

Characterization of the tunicamycin gene cluster unveiling unique steps involved in its biosynthesis

Tunicamycin,a potent reversible translocase I inhibitor,is produced by several Actinomycetes species.The tunicamycin structure is highly unusual,and contains an 11-carbon dialdose sugar and anα,β-1″,11′-glycosidic linkage.Here we report the identification of a gene cluster essential for tunicamycin biosynthesis by high-throughput heterologous expression(HHE)strategy combined with a bioassay.Introduction of the genes into heterologous non-producing Streptomyces hosts results in production of tunicamycin by these strains,demonstrating the role of the genes for the biosynthesis of tunicamycins.Gene disruption experiments coupled with bioinformatic analysis revealed that the tunicamycin gene cluster is minimally composed of 12 genes(tunA–tunL).Amongst these is a putative radical SAM enzyme(Tun B)with a potentially unique role in biosynthetic carbon-carbon bond formation.Hence,a seven-step novel pathway is proposed for tunicamycin biosynthesis.Moreover,two gene clusters for the potential biosynthesis of tunicamycin-like antibiotics were also identified in Streptomyces clavuligerus ATCC 27064 and Actinosynnema mirums DSM 43827.These data provide clarification of the novel mechanisms for tunicamycin biosynthesis,and for the generation of new-designer tunicamycin analogs with selective/enhanced bioactivity via combinatorial biosynthesis strategies.