Bleomycin-induced gene expression in E.coli carrying blmA gene

博莱霉素诱导携带 blmA 基因的大肠杆菌中的基因表达

• 批准号: 05454570
• 负责人: SUGIYAMA Masanori
• 金额: $ 4.42万
• 依托单位: HIROSIMA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1993
• 资助国家: 日本
• 起止时间: 1993 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05454570/
• 关键词: Bleomycim; Inducer; beta-lactamase; Antitumor agent; インデューサー

A bleomycin (Bm)-producing microorganism, Streptomyces verticillus ATCC15003 possesses two Bm resistance genes designated blmA and blmB.When one of these genes, designatedblmA,was inserted into pUC18 (or pUC19) plasmid and introduced into Escherichia coli TG1, the transformed cells expressed high level-resistance to Bm. In the present study we show that E.coli harboring the blmA-encoding pUC vector overproduced a 30-kDa protein when cultured in the presence of Bm. The Bm-induced protein was identified as the beta-lactamase encoded by the ampicillin resistance gene on the plasmid. In addition, we found that although the Bm family such as pepleomycin, phleomycin, cleomycin and liblomycin gave rise to beta-lactamase induction, bleomycininc acid which lacks terminal amine moiety in Bm molecule, did scarcely. We propose a hypothesis that Bm, an antibiotic inhibitor of DNA synthesis, acts as a an inducer (or an activator) for the expression of a specific gene in the presence of blmA.On the other hand, we constructed a chimeric plasmid designatd as intelligent vector pMAB70, which carried blmA and a tyrosinase structual gene put under the control of beta-lactamase promoter. E.coli harboring pMAB70 produced the melanine pigment, when cultured in the presence of Bm.

轮枝链霉菌ATCC 15003具有两个抗博莱霉素基因blmA和blmB，将其中一个基因blmA插入pUC 18（或pUC 19）质粒中，转化大肠杆菌TG 1，获得了高水平的抗博莱霉素的转化细胞。在本研究中，我们表明，大肠杆菌窝藏blmA编码的pUC载体过度产生30 kDa的蛋白质时，在Bm的存在下培养。经鉴定，该细菌诱导蛋白为质粒上氨苄青霉素抗性基因编码的β-内酰胺酶。此外，我们发现，虽然Bm家族如培洛霉素、腐草霉素、洁霉素和利布霉素引起β-内酰胺酶的诱导，但Bm分子中缺少末端胺的博莱霉素酸几乎不引起β-内酰胺酶的诱导。我们提出了DNA合成的抗生素抑制剂Bm在blmA的存在下作为特定基因表达的诱导剂（或激活剂）的假说，并构建了一个含有blmA和β-内酰胺酶启动子控制下的酪氨酸酶结构基因的嵌合质粒pMAB 70。当在Bm存在下培养时，携带pMAB 70的大肠杆菌产生黑色素。

项目成果

Sugiyama,M.: "Reconbinant Microbes for Industrial and Agricultural Applications" Marcel Dekker,Inc.(Murooka,Y.and Imanaka,T.eds), 875 (1994)

Sugiyama,M.：“用于工业和农业应用的重组微生物”Marcel Dekker,Inc.（Murooka,Y. 和 Imanaka,T.eds），875 (1994)

Sugiyama,M.: "Inactivation of bleomycin by an N-acetyltransferase in the bleomycin-producing strain Streptomyces verticillus" FEMS Microbiology Letters. 121. 81-86 (1994)

Sugiyama,M.：“博来霉素生产菌株轮枝链霉菌中的 N-乙酰转移酶使博莱霉素失活”FEMS 微生物学快报。

Sugiyama M.: "Characterization by molecular cloning of two genes from Streptomyces verticillus encoding resistance to bleomycin" Gene. 151. 11-16 (1994)

Sugiyama M.：“通过分子克隆对来自轮枝链霉菌的编码博莱霉素抗性的两个基因进行表征”基因。

Sugiyama,M.: "Inactivation of bleomycin by an N‐acetyltransferase in the bleomycin‐producing strain Streptomyces verticillus" FEMS Microbiology Letters. 121. 81-86 (1994)

Sugiyama, M.：“产博莱霉素菌株链霉菌中 N-乙酰转移酶对博来霉素的灭活”FEMS 微生物学快报。 121. 81-86 (1994)

Sugiyama,M.: "Recombinant Microbes for Industrial and Agricultural Applications" Marcel Dekker Inc.(Murooka Y.and Imanaka,T.eds), 875 (1994)

Sugiyama,M.：“用于工业和农业应用的重组微生物”Marcel Dekker Inc.（Murooka Y. 和 Imanaka,T.eds），875 (1994)