Effect of amino acid substitutions in conserved residues in the leader peptide on biosynthesis of the lantibiotic mutacin II

The lantibiotic mutacin II, produced by Streptococcus mutans T8, is a ribosomally synthesized peptide antibiotic that contains thioether amino acids such as lanthionine and methyllanthionine as a result of post-translational modifications. The mutacin II leader peptide sequence shares a number of identical amino acid residues with class AII lantibiotic leader peptides. To study the role of these conservative residues in the production of active antimicrobial mutacin, 15 mutations were generated by site-directed mutagenesis. The effects of these substitutions vary from no effect to complete block-out. Mutations G-1A, G-2A, I-4D, and L-7K completely blocked the production of mature mutacin. Other mutations (I-4V, L-7M, E-8D, S-11T/A, V-12I/A, and E-13D) had no detectable effect on mutacin production. The changes of Glu-8 to Lys, Val-12 to Leu, Glu-13 to Lys reduced the mutacin production level to about 75%, 50%, and 10% of the wild-type, respectively. Thus, our data indicated that some of these conserved residues are essential for the mutacin biosynthesis, whereas others are important for optimal biosynthesis rates.     

变链素活性与变形链球菌基因多态性的关系研究

目的探讨变链素的活性与变形链球菌(MS)基因多态性的关系。方法在AP-PCR基因分型的基础上,选择来自单基因型定植的个体50株MS为单基因型组,另50株来自多基因型共同定植的个体为多基因型组,用平板法检测2组菌株产生变链素对10个指示株的抑制情况,T-检验比较2组菌株抑菌环和抑菌谱的均数差异。结果所有的实验株(100%)均可产生抑制6~8个指示株的变链素,抑菌环和抑菌谱在不同个体之间变异,组间均数的比较不具有显著性(P值分别是0.12,1.79)。结论多基因型MS定植的口腔,在产生变链素方面似乎不具有优势。     

液体培养基中天然变链素的纯化

目的 从变形链球菌临床株的液体培养基中分离纯化变链素,为进一步从分子水平研究变链素奠定基础.方法 通过抑菌活性检测,从变链临床株中选择出抑菌活性较强的菌株.用氯仿抽提法从该菌株的培养液中粗提变链素,经固相萃取和反相高效液相色谱(RP-HPLC)对粗提物进行纯化.结果 获得变链素活性较强的菌株"1G".从其200 ml液体培养基中粗提出变链素约15 μg,经固相萃取柱洗脱,再经过RP-HPLC 2次纯化,得到有抑菌活性的成分,此为纯化的变链素.结论 变链素分子量小,分离提纯步骤复杂,本实验得到纯化的变链素,为下一步研究变链素的氨基酸序列和基因序列奠定了基础.     

Streptococcus mutans strain N produces a novel low molecular mass non-lantibiotic bacteriocin

Streptococcus mutans strain N was shown to have bacteriocin production and immunity characteristics consistent with those of Group I mutacin-producing strains of S. mutans. The bacteriocin mutacin N was purified from agar cultures of S. mutans strain N using XAD andp6 reversed phase chromatography. The molecular mass of mutacin N was 4806 Da and the entire 49 amino acid sequence was determined by N-terminal sequencing. Database searches indicate that mutacin N is a novel bacteriocin, but with some homology to the protein IIC domain of a hypothetical sugar-phosphotransferase enzyme from Acholeplasma florum.