GROWTH AND METABOLISM OF ORAL MICROORGANISMS

口腔微生物的生长和代谢

• 批准号: 5201774
• 负责人: S A ROBRISH
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF DENTAL & CRANIOFACIAL RESEARCH
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/5201774
• 关键词: Escherichia coli; Fusobacterium; Streptococcus mutans; antibody; carbohydrate metabolism; carbohydrate transport; dithiol; enzyme activity; genetic strain; hydro lyase; hydrolysis; maltose; microorganism culture; microorganism growth; microorganism metabolism; molecular cloning; nucleic acid sequence; oral bacteria; phosphoenolpyruvate; phosphotransferases; protein purification; protein sequence

Activity for the hydrolysis of maltose 6 phosphate [maltose 6P] has been stabilized in sonic extracts of maltose grown Fusobacterium mortiferum (ATCC 25557) using dithiothreitol [DTT]. All of the activity was found in a single protein with a molecular weight of 49 KD following purification in the presence of DTT and assay with an artificial alpha glucoside phosphate. The purified protein hydrolyzed authentic maltose 6P to equimolar amounts of glucose 6P and glucose confirming a phosphoenolpyruavate phosphotransferase (PTS) activity for maltose use by F. mortiferum. The hydrolytic activity of maltose 6P hydrolase was restricted to alpha glucoside phosphates and required Mn++. An artificial alpha glucoside phosphate proved necessary for the purification of the protein and an antibody was made to the resultant 49 KD homogeneous protein with enzyme activity. Maltose 6P hydrolase activity was induced in F. mortiferum by growth on a variety of sugars; mostly, but not exclusively, alpha glucosides. The specific activity of the hydrolase enzyme was proportional to a hierarchy of growth sugars and the activity was exclusively in a 49 KD protein. The sequence of 32 amino acids from the NH2 terminal end of the maltose 6P hydrolase was determined and the activity was cloned, sequenced, and expressed in Escherichia coli. Growth on beta glucosides induced the anaerobic use of cellobiose by washed cell suspensions of F. mortiferum. A 54 KD protein with activity for the hydrolysis of a beta glucoside phosphate has been purified and identified from sonic extracts of cellobiose grown cells. The properties of this beta glucoside phosphate hydrolase and its functioning in a cellobiose PTS in F. mortiferum are now being studied. Maltose is used by F. necrophorum, but by a different mechanism than that reported by us for F. mortiferum. The 49 KD protein for the maltose 6P hydrolase could not be demonstrated in a sonic extract of maltose grown cells of F. necrophorum. Two strains of Streptococcus mutans, reported to have a maltose PTS, had no detectable activity when sonic extracts were tested with antibody to the F. mortiferum maltose 6P hydrolase.

已经测定了麦芽糖6磷酸[麦芽糖6P]的水解活性。 在麦芽糖生长的死亡梭杆菌的声波提取物中稳定 (ATCC 25557）。 所有的活动都是在 纯化后得到分子量为49 KD的单一蛋白 在DTT存在下，用人工α-葡糖苷进行测定 磷酸盐 纯化的蛋白质水解真正的麦芽糖6P， 等摩尔量的葡萄糖6P和葡萄糖证实了 磷酸烯醇化氨基酸磷酸转移酶（PTS）活性，用于麦芽糖的使用， F.死亡 麦芽糖6P水解酶的水解活性为 限于α-葡糖苷磷酸盐，并且需要Mn ++。 人工 证明α-葡萄糖苷磷酸对于纯化是必要的 蛋白质，并对所得的49KD均一的蛋白质制备抗体 具有酶活性的蛋白质。 麦芽糖6P水解酶活性被诱导 in F. mortiferum通过生长在各种糖上;大多数情况下，但不是 仅限于α-葡糖苷。 水解酶的比活性 酶与生长糖的层次成正比， 仅存在于49KD蛋白中。 来自大肠杆菌的32个氨基酸的序列 测定麦芽糖6P水解酶的NH 2末端， 活性进行克隆、测序，并在大肠杆菌中表达。 增长 对洗涤细胞诱导纤维二糖厌氧利用的β-葡萄糖苷的影响 悬浮液F.死亡 一种54KD的蛋白，其具有对 已经纯化和鉴定了β-葡糖苷磷酸的水解 从纤维二糖生长的细胞的声波提取物。 这个的属性 β-葡萄糖苷磷酸水解酶及其在纤维二糖PTS中的功能 in F.目前正在研究中。 麦芽糖被F. 坏死，但其机制与我们报道的F. 死亡 麦芽糖6P水解酶的49 KD蛋白不能被纯化。 在麦芽糖生长的F.坏死。 据报道，有两种变形链球菌具有麦芽糖PTS， 当声波提取物用抗体测试时， 食品药品管理桑椹麦芽糖6P水解酶