1. 吉林大学超分子结构与材料国家重点实验室，吉林 长春 130012 2. Université de Lyon, Villeurbanne, F-69622, France; CNRS UMR 5246 ICBMS, Villeurbanne, F-69622, France
Kinetic Study on Inhibition Effects of Dansyl-L-Phenylalanine and L-Phenylalanine on Calf Intestinal Alkaline phosphatase
LI Li-na1, 2, WU Yu-qing1*, René Buchet2
1. State Key Laboratory for Supramolecular Structure and Materials, Jilin University, Changchun 130012, China 2. Université de Lyon, Villeurbanne, F-69622, France; CNRS UMR 5246 ICBMS, Villeurbanne, F-69622, France
Abstract：To evaluate the inhibition effect of dansyl-L-phenylalanine on calf intestinal alkaline phosphatase (CIAP), UV-Vis spectrophotometric method was employed. It was found that dansyl-L-phenylalanine can selectively inhibit CIAP. The kinetic inhibition processes of dansyl-L-phenylalanine and L-phenylalanine were comparatively studied. The authors’ finding elucidates that at the optimized alkaline pH of alkaline phosphatase (pH 10.4) and 37 ℃, dansyl-L-phenylalanine can inhibit alkaline phosphatase activity of CIAP efficiently and specifically, similar as L-phenylalanine. Both inhibition types were uncompetitive inhibition resulting from the double reciprocal curve fitting of v versus substrate concentrations, and the inhibition constants Ki of both inhibitors were determined to be 2.3 and 1.1 mmol·L-1 respectively, both of which were at millimolar level. The investigation of the inhibition effect of dansyl modified L-phenylalanine on calf intestinal alkaline phosphatase not only helped get insight into the detailed inhibition mechanism of L-phenylalanine on tissue specific alkaline phosphatase, such as in the case of intestinal alkaline phosphatase, but also provided the possibility to employ fluorescence spectroscopy by labeling the specific inhibitors of alkaline phosphatase with chromophoric groups.
李丽娜1, 2, 吴玉清1*, René Buchet2. 丹酰-L-苯丙氨酸和L-苯丙氨酸对小牛肠碱性磷酸酶抑制作用动力学的比较性研究[J]. 光谱学与光谱分析, 2009, 29(10): 2820-2823.
LI Li-na1, 2, WU Yu-qing1*, René Buchet2. Kinetic Study on Inhibition Effects of Dansyl-L-Phenylalanine and L-Phenylalanine on Calf Intestinal Alkaline phosphatase. SPECTROSCOPY AND SPECTRAL ANALYSIS, 2009, 29(10): 2820-2823.
[1] Schwartz J H, Lipmann F. Proc. Nat. Acad. Sci. USA, 1961, 47: 1996. [2] Zhang L, Balcerzak M, Radisson J, et al. Journal of Biological Chemistry, 2005, 280: 37289. [3] Harris H. Clin. Chim. Acta, 1989, 186: 133. [4] Noda M, Yoon K, Rodan G A, et al. J. Cell Biol., 1987, 105: 1671. [5] Einhorn T A, Gordon S L, Siegel S A, et al. J. Orthop. Res., 1985, 3: 160. [6] Denier C, Brisson-Lougarre A, Biasini G, et al. BMC Biochemistry, 2002, 3: 2. [7] Kozlenkov A, Du M H, Cuniasse P, et al. J. Bone. Miner. Res., 2004, 19: 1862. [8] LI Zhong-qin, XU Xiao-ping, WANG Wu(李忠琴, 许小平, 王 武). Spectroscopy and Spectral Analysis(光谱学与光谱分析), 2008, 28(9): 2169. [9] Fishman W H, Sie, H G. Enzymologia, 1971, 41: 140. [10] Lin C W, Sie H G, Fishman W H. Biochem. J., 1971, 124: 509. [11] Baltas M, Lapeyre C, Bedos-Belval F, et al. Plant Physiology and Biochemistry, 2005, 43: 746. [12] Hoylaerts M F, Manes T, Millán J L. Biochem J., 1992, 286: 23. [13] Kozlenkov A, Manes T, Hoylaerts M F, et al. J. Biol. Chem., 2002, 277(25): 22992.
