摘要
本文研究高粱外种皮原花青素四聚体(sorghum episperm procyanidin tetramer,SEPT)对远缘链球菌6715(S.sobrinus 6715)初始粘附及其致龋毒力因子-葡萄糖基转移酶(glucosyltransferase,GTF)的抑制效果。采用唾液包被羟基磷灰石(saliva coated hydroxylapatite,SHA)体外模型研究SEPT对S.sobrinus 6715初始粘附的影响。通过PCR扩增S.sobrinus 6715 GTF-I的催化活性区(catalytic region,CAT)的基因片段到质粒Pgex-4T-1中,转化至大肠杆菌T7 Express competent,IPTG诱导表达,纯化浓缩表达产物,获得目标蛋白GTF-I/CAT。再用不同浓度SEPT处理GTF-I/CAT,根据其水不溶性多糖产量(water insoluble glucosyltransferase,WIS)评价SEPT对GTF-I/CAT酶活力的抑制。实验结果表明,SEPT能够有效抑制S.sobrinus 6715在SHA的初始粘附,并具有一定的浓度效应关系;通过基因克隆表达获得的GTF-I/CAT蛋白酶活为8.446 m IU,产多糖能力为1.603 mg/(mg·h)。SEPT能明显抑制GTF-I/CAT产生WIS(p<0.01)。说明SEPT既可以有效抑制S.sobrinus 6715的初始粘附过程,又可以通过作用GTF-I的CAT区减少WIG的合成,抑制蔗糖依赖性粘附减少S.sobrinus 6715在牙齿表面的粘附聚集,从而干预龋齿的发生。
The inhibitory effect of sorghum episperm procyanidin tetramer（SEPT） on the initial adherence of Streptococcus sobrinus 6715 and its cariogenic factor glucosyltransferase（GTF） was investigated.The saliva-coated hydroxyapatite（SHA） was used as an in vitro model to measure the inhibitory effect of SEPT on the initial adherence of S.sobrinus 6715.The GTF-I catalytic region（CAT） gene of S.sobrinus 6715 was amplified by polymerase chain reaction（PCR）,cloned into a Pgex-4T-1 vector,and transformed into T7 Express competent of Escherichia coli.The protein expression was induced by isopropyl β-D-1-thiogalactopyranoside（IPTG）,and the expressed product was purified and concentrated to obtain the GTF-I/CAT target protein.Subsequently,different concentrations of SEPT samples were used to treat GTF-I/CAT,and the inhibitory effect of SEPT on GTF-I/CAT enzyme activity was determined based on the amount of water-insoluble glucan（WIG） formed by GTF-I/CAT.The results showed that SEPT could effectively decrease the initial adherence of S.sobrinus 6715 to SHA in a dose-dependent manner.The enzyme activity of GTF-I/CAT obtained from the gene cloning and expression was 8.446 m IU and the WIG formation ability was 1.603 mg/（mg·h）.SEPT could effectively suppress the formation of WIG by acting on GTF-I/CAT（P〈0.01）.The results demonstrated that SEPT was effective in inhibiting the initial adherence of S.sobrinus 6715 to SHA and reducing WIG synthesis by acting on the CAT region of GTF-I.Therefore,SEPT can decrease the adhesion and aggregation of S.sobrinus 6715 on tooth surface and inhibit the sucrose-dependent adherence,thereby preventing the occurrence of dental caries.
作者
田晶
鲁群
鲁亮
张永霞
刘睿
TIAN Jing LU Qun LU Liang ZHANG Yong-xia LIU Rui(Key Laboratory of Environment Correlative Dietology, College of Food Science and Technology, Huazhong Agricultural University, Wuhan 430070, China PuJian Biological Technology Co., LTD., Wuhan 430070, China)
出处
《现代食品科技》
EI
CAS
北大核心
2017年第2期48-54,共7页
Modern Food Science and Technology
基金
国家自然基金项目(31271939)
