羊栖菜多糖提取物对酪氨酸酶的抑制作用研究
详细信息Study on inhibition effect of Sargassum fusiforme polysaccharide extracts on activity of tyrosinase
-
摘要: 目的:研究羊栖菜多糖提取物对酪氨酸酶活性的抑制作用和对结构的影响。方法:提取羊栖菜多糖并测定含量,以L-DOPA为底物,在475nm处测定吸光度,观察羊栖菜多糖提取物对酪氨酸酶二酚酶活力的影响;测定羊栖菜多糖作用下酪氨酸酶内源荧光和ANS结合的荧光的变化,以判断对酶结构的影响情况。结果:羊栖菜多糖提取物对蘑菇酪氨酸酶有明显的抑制作用,可以导致酪氨酸酶的二酚酶活力下降。羊栖菜多糖使二酚酶活力下降一半时的抑制剂浓度（IC50）为（2.16±0.37）mg/mL。羊栖菜多糖对酪氨酸酶的抑制作用是一个可逆过程,抑制类型为混合型抑制。通过动力学分析,测得羊栖菜多糖对酪氨酸酶的抑制常数（Ki）为（4.34±0.56）mg/mL。荧光测定结果表明,羊栖菜多糖的结合引起酪氨酸酶三级结构的明显改变。结论:羊栖菜多糖与酪氨酸酶的结合引起酶结构的变化,并以混合抑制的方式可逆抑制酪氨酸酶的二酚酶活性。本实验为进一步研究和设计新型酪氨酸酶抑制剂奠定科学基础。Abstract: Objective:To study the inhibitory effect of the Sargassum fusiforme polysaccharide (SFPS) extracts on the activity of tyrosinase for diphenolase.Methods:Extract SFPS and measure the content of polysaccharide.To investigate the effect of SFPS extracts on the diphenolase activity, the absorbance of the solution containing several concentrations of the SFPS extracts was measured at 475nm, using the L-DOPA as the substrate.Endogenous fluorescence and ANS-binding fluorescence of tyrosinase were tested with different concentrations of SFPS.Results:SFPS could obviously inhibit the activity of tyrosinase for diphenolsae.The inhibitory concentration leading to 50% activity lost (IC 50) was estimated to be (2.16±0.37) mg/mL.The results showed that SFPS was a reversible inhibitor of mushroom tyrosinase.Lineweaver-Burk plots indicated that SFPS was a mixed-type inhibitor, and its inhibition constant Ki were (4.34±0.56) mg/mL.Endogenous fluorescence and ANSbinding fluorescence test of tyrosinase indicated that SFPS combined with the tyrosinase caused significant changes in the structure.Conclusion:SFPS combined with the tyrosinase caused significant changes in the structure and reversible inhibited the activity of tyrosinase for diphenolsae in a mixed-type manner.Together these results provided a scientific basis for the design and development of new inhibitors in the future.
-
[1] Krner A M, Pawelek J M.Mammalian tyrosinase catalyzes three reaction in the biosynthesis of melanin[J].Science, 1982, 217, 4565:1163-1165.
[2] Sánchez-Ferrer A, Rodriguez-López J N, Garcia-Cánovas F, et al.Tyrosinase:a comprehensive review of its mechanism[J].Biochim Biophys Acta, 1995, 1247 (1) :1-11.
[3] Tripathi R K, Hearing V J, Urabe K, et al.Mutational mapping of the catalytic activities of human tyrosinase[J].J Biol Chem, 1992, 267, 33:23707-23712.
[4] Petrescu S M, Popescu C I, Petrescu A J, et al.The glycosylation of tyrosinase in melanoma cells and the effect on antigen presentation[J].Adv Exp Med Biol, 2003, 535:257-269.
[5] Zou X W, Jiang Z S.Advances in studies on botanical inhibitors of tyrosinase[J].China Tradit Herb Drug, 2004, 35 (6) :702-705.
[6] Karioti A, Protopappa A, Megoulas N, et al.Identification of tyrosinase inhibitors from Marrubium velutinum and Marrubium cylleneum[J].Bioorgan Med Chem, 2007, 15 (7) :2708-2714.
[7] 洪咏平, 戴志远.羊栖菜食品的开发和利用[J].中国水产, 2002 (1) :75-76. [8] 武晓丹, 邹翔.海洋藻类多糖的药理研究[J].哈尔滨商业大学学报:自然科学版, 2005, 21 (2) :137-138. [9] 周峙苗.羊栖莱多糖的提取和纯化研究[D].杭州:浙江工业大学, 2004. [10] 汲晨锋, 季宇彬, 王翀, 等.羊栖菜多糖含量测定及多糖组分分析[J].世界科学技术-中医药现代化, 2006, 5 (8) :49-51. [11] 张惟杰.糖复合物生化研究技术[M].杭州:浙江大学出版社, 2003, 1-348. [12] Park Y D, Kim S Y, Lyou Y J, et al.A new type of uncompetitive inhibition of tyrosinase induced by Cl-binding[J].Biochimie, 2005, 87 (11) :931-937.
[13] Han H Y, Lee J R, Xu W A, et al.Effect of Cl-on tyrosinase:complex inhibition kinetics and biochemical implication[J].J Biomol Struct Dyn, 2007, 25 (2) :165-171.
[14] Passi S, Nazzaro-Porro M, Molecular basis of substrate and inhibitory specificity of tyrosinase:phenolic compounds[J].Br J Dermatol, 1981, 104 (6) :659-665.
[15] Cabanes J, Garcia-Canovas F, Tudela J, et al.L-mimosine a slow-binding inhibitor of mushroom tyrosinase[J].Phytochemistry, 1987, 26 (4) :917-919.
[16] Klabunde T, Eicken C, Sacchettini J C, et al.Crystal structure of a plant catechol oxidase containing a dicopper center[J].Nat Struct Biol, 1998, 5 (12) :1084-1090.
[17] Pérez-Gilabert M, García-Carmona F.Dimethyl sulfide, a volatile flavor constituent, is a slow-binding inhibitor of tyrosinase[J].Biochem Biophys Res Commun, 2001, 285 (2) :257-261.
计量
- 文章访问数: 125
- HTML全文浏览量: 12
- PDF下载量: 714
下载:
