Isolation of a-1,3-Glucanase-Producing Strains

• Main Authors: Hsiao-Yi Zu, 朱孝益
• Other Authors: Meng-Hsiao Meng
• Format: Others
• Language: zh-TW
• Published: 1999
• Online Access: http://ndltd.ncl.edu.tw/handle/84279320609978359893

碩士 === 國立中興大學 === 農業生物科技學研究所 === 87 === According to dental research, the reason for caries and denudina is dental plaque. Bacterial plaque is a very sticky material that absorbs caries bacteria on the surface of tooth. These caries bacteria can continuously produce acid. The acid environment will damage enamel of teeth. On the other hand, bacteria plaques are also absorb some cation to become teeth-stone. The indurative teeth-stone accumulates between gingiva and tooth. For a long term, worsen situation brings gingivitis and denudation of the teeth. Dental plaque is the film of micro organisms found on the tooth surface embedded in a matrix of the polymers of salivary and bacterial origin. (J Dent Res July 1992 Vol. 71 No. 71431-1438) The main component of bacteria plaque is a-1,3-glucan. Glucosyltransferase (Gtf) which come from Streptococcus spp. catalyzes sucrose become a-1,3-glucan. The goal of my study is to find bacteria which can decompose a-1,3-glucan, then I will purify the enzyme from these bacteria and analyse it's enzyme kinetic. According to the previous reports, Streptococcus mutans OMZ 176, which is important in formation of dental plaque(J. Bactreiol vol 124 1975 1488-1501), however it could not be obtained from bacteria stock center. So we used S. sobrinus ATCC 14757, that has high similarity with S. mutans OMZ 176, to produce a-1,3-glucan. In order to get high quality substrate (a-1,3-glucan), we clone gtf gene from S. sobrinus ATCC 14757. Transformation this gene to E. coli. by pUC18 vector. We hope to get pure and a large of enzyme to produce high quality a-1,3-glucan. Meanwhile, we extracted secretion protein from S. sobrinus ATCC 14757 blood culture medium by ammonium sulfate precipitate method. In 35%-45% saturated ammonium sulfate concentration, we got a group of protein which could produce insoluble carbohydrate (like dental plaque component) from sucrose. We collected soil specimens from fifteen areas and get three hundred forty seven different soil specimens in Taiwain. By enrichment culture of soil specimens, used insoluble carbohydrate as sole carbon source, forty-three organisms could survival. Next step, I subcultured these bacteria by cell-wall-glucan of A niger. Isolate M15 and M34 were found to grow better than the other organisms. But the released glucose activity did not be detected. I hope to improve the enzyme to commercialize, so everyone can use it conveniently and have adequate oral hygiene.