Tognoni A, Carrera P, Galli G, Lucchese G, Camerini B, Grandi G
Department of Molecular Biology, Via San, Milan, Italy.
J Gen Microbiol. 1989 Jan;135(1):37-45. doi: 10.1099/00221287-135-1-37.
A strain of Pseudomonas sp., SMP1, isolated from a soil sample collected in the Monterotondo area (Rome), secreted isoamylase activity into the culture medium. The enzyme was purified and optimal reaction and stability conditions were determined by varying pH and temperature. The chemico-physical properties of the enzyme were similar to those of the isoamylase purified in Japan more than 20 years ago from 'Pseudomonas amyloderamosa' strain SB15. A genomic library of SMP1 was prepared in Escherichia coli using pUC12 as vector. Two isoamylase-producing colonies were identified out of 6300 screened. The hybrid plasmids isolated from the two clones showed common restriction patterns. The chromosomal portion of one of these plasmids (pSM257) was completely sequenced. Comparison between the deduced amino acid sequence of the isoamylase and the published sequences of other amylolytic enzymes showed the presence of conserved domains.
从罗马蒙特罗通多地区采集的土壤样本中分离出的一株假单胞菌属菌株SMP1，能向培养基中分泌异淀粉酶活性。该酶经过纯化，并通过改变pH值和温度确定了最佳反应和稳定性条件。该酶的化学物理性质与20多年前在日本从“解淀粉假单胞菌”菌株SB15中纯化的异淀粉酶相似。以pUC12为载体，在大肠杆菌中构建了SMP1的基因组文库。在筛选的6300个菌落中鉴定出两个产生异淀粉酶的菌落。从这两个克隆中分离出的杂交质粒显示出共同的限制性酶切图谱。对其中一个质粒（pSM257）的染色体部分进行了全序列测定。异淀粉酶推导的氨基酸序列与其他淀粉分解酶已发表序列的比较显示存在保守结构域。
