Klotho-related Protein Is a Novel Cytosolic Neutral β-Glycosylceramidase
作者： Yasuhiro Hayashi , Nozomu Okino , Yoshimitsu Kakuta , Toshihide Shikanai , Motohiro Tani
关键词: Enzyme kinetics 、 Cytosol 、 Recombinant DNA 、 Biology 、 Biochemistry 、 Glycosylceramidase 、 Klotho 、 Small interfering RNA 、 TIM barrel 、 Molecular biology 、 Enzyme
摘要: Using C6-NBD-glucosylceramide (GlcCer) as a substrate, we detected the activity of conduritol B epoxide-insensitive neutral glycosylceramidase in cytosolic fractions zebrafish embryos, mouse and rat brains, human fibroblasts. The candidates for enzyme were assigned to Klotho (KL), whose family members share β-glucosidase-like domain but natural substrates are unknown. Among this family, only KL-related protein (KLrP) is capable degrading C6-NBD-GlcCer when expressed CHOP cells, which Myc-tagged KLrP was exclusively distributed cytosol. In addition, knockdown endogenous by small interfering RNA increased cellular level GlcCer. purified recombinant hydrolyzed 4-methylumbelliferyl-glucose, C6-NBD-GlcCer, authentic GlcCer at pH 6.0. also corresponding galactosyl derivatives, each kcat/Km much lower than that glucosyl derivatives. x-ray structure 1.6A resolution revealed (β/α)8 TIM barrel, Glu165 Glu373 carboxyl termini β-strands 4 7 could function an acid/base catalyst nucleophile, respectively. substrate-binding cleft occupied with palmitic acid oleic crystallized complex glucose. found fit well crystal KLrP. Collectively, identified be involved novel nonlysosomal catabolic pathway
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A H Futerman, R E Pagano, Determination of the intracellular sites and topology of glucosylceramide synthesis in rat liver Biochemical Journal. ,vol. 280, pp. 295- 302 ,(1991) , 10.1042/BJ2800295
R Charles Scriver, The Metabolic and Molecular Bases of Inherited Disease ,(1995)
L W Cary, T A Koerner, S C Li, Y T Li, Carbon 13 NMR spectroscopy of a cerebroside. Proof of the beta-pyranosyl structure of D-glucosylceramide. Journal of Biological Chemistry. ,vol. 254, pp. 2326- 2328 ,(1979) , 10.1016/S0021-9258(17)30224-7
William S HAYS, Steven A. JENISON, Takashi YAMADA, Andrzej PASTUSZYN, Robert H. GLEW, Primary structure of the cytosolic beta-glucosidase of guinea pig liver. Biochemical Journal. ,vol. 319, pp. 829- 837 ,(1996) , 10.1042/BJ3190829
Heidrun Matern, Henrike Boermans, Siegfried Matern, Friedrich Lottspeich, Molecular Cloning and Expression of Human Bile Acid β-Glucosidase Journal of Biological Chemistry. ,vol. 276, pp. 37929- 37933 ,(2001) , 10.1074/JBC.M104290200
Anastassis Perrakis, Richard Morris, Victor S. Lamzin, Automated protein model building combined with iterative structure refinement. Nature Structural & Molecular Biology. ,vol. 6, pp. 458- 463 ,(1999) , 10.1038/8263
T. Kobayashi, K. Suzuki, The glycosylceramidase in the murine intestine. Purification and substrate specificity. Journal of Biological Chemistry. ,vol. 256, pp. 7768- 7773 ,(1981) , 10.1016/S0021-9258(18)43343-1
Zbyszek Otwinowski, Wladek Minor, Processing of X-ray diffraction data collected in oscillation mode Methods in Enzymology. ,vol. 276, pp. 307- 326 ,(1997) , 10.1016/S0076-6879(97)76066-X
L.B. Daniels, P.J. Coyle, Y.B. Chiao, R.H. Glew, R.S. Labow, Purification and characterization of a cytosolic broad specificity beta-glucosidase from human liver. Journal of Biological Chemistry. ,vol. 256, pp. 13004- 13013 ,(1981) , 10.1016/S0021-9258(18)42997-3
Vasantha L. Kolachala, Tracy Sitler Obertone, Lixin Wang, Didier Merlin, Shanthi V. Sitaraman, Adenosine 2b receptor (A2bR) signals through adenylate cyclase (AC) 6 isoform in the intestinal epithelial cells. Biochimica et Biophysica Acta. ,vol. 1760, pp. 1102- 1108 ,(2006) , 10.1016/J.BBAGEN.2006.03.010