Entrapping enzyme in a functionalized nanoporous support

Journal of the American Chemical Society

Volumn 124, Issue 38, 2002, Pages 11242-11243

  Lei, Chenghong ^a   Shin, Yongsoon ^a   Liu, Jun ^b   Ackerman, Eric J ^a  

^a PACIFIC NORTHWEST NATIONAL LABORATORY   (United States)

^b SANDIA NATIONAL LABORATORIES   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

NANOPARTICLE;

ARTICLE; BINDING KINETICS; COMPARATIVE STUDY; ENZYME IMMOBILIZATION; ENZYME STABILITY; ENZYME STRUCTURE; MOLECULAR BIOLOGY; PH; PHYSICAL CHEMISTRY; POROSITY; SURFACE PROPERTY;

ARYLDIALKYLPHOSPHATASE; ENZYME STABILITY; ENZYMES, IMMOBILIZED; ESTERASES; MICROSCOPY, ELECTRON; PEROXIDES; SILICON DIOXIDE;

EID: 0037174353     PISSN: 00027863     EISSN: None     Source Type: Journal    
DOI: 10.1021/ja026855o     Document Type: Article

References (19)

1. Bickerstaff, G. F. (Ed.) Immobilization of Enzymes and Cells; Humana Press: Totowa, NJ, 1997.
2. Livage, J.; Coradin, T.; Roux, C. J. Phys. Condes. Matter. 2001, 13, R673-691.
3. (a) Braun, S.; Rappoport, S.; Zusman, R. Mater. Lett. 1990, 10, 1-5.
4. (b) Wei, Y.; Xu, J.; Feng, Q.; Dong, H.; Lin, M. Mater. Lett. 2000, 44, 6-11.
5. (a) Feng, X.; Fryxell, G. E.; Wang, L.-Q.; Kim, A. Y.; Liu, J.; Kemner, K. M. Science 1997, 276, 923-926.
6. (b) Liu, J.; Feng, X.; Fryxell, G. E.; Wang, L.-Q.; Kim, A. Y.; Gong, M. Adv. Mater 1998, 10, 161-165.
7. Yiu, H. H. P.; Wright, P. A.; Botting, N. P. Microporous Mesoporous Mater. 2001, 44-45, 763-768.
8. Yiu, H. H. P.; Wright, P. A.; Botting, N. P. J. Mol. Catal. B: Enzym. 2001, 15, 81-92.
9. Zhao, D.; Feng, J.; Huo, Q.; Melosh, N.; Fredrickson, G. H.; Chmelka, B. F.; Stucky, G. D. Science 1998, 279, 548-552.
10. Linsen, B. G.; van den Heuvel, A. The Solid Gas Interface; Flood, E. A., Ed.; Mercel Dekker: New York, 1967; Vol. 2, p 1025.
11. Normal porous silica is uncoated silica bulk material with a pore size of 30 nm with a 12-μm bead size from PolyLC Inc., USA, item no. BMSI 1203.
12. 4
13. (a) Singh, A. K.; Flounders, A. W.; Volponi, J. V.; Ashley, C. S.; Walley, K.; Schoeniger, J. S. Biosens. Bioelectron. 1999, 98, 703-713.
14. (b) LeJeune, K. E.; Wild, J. R.; Russel, A. J. Nature 1998, 395, 27 -28.
15. (c) LeJeune, K. E.; Russell, A. J. Biotechnol. Bioeng. 1996, 51, 450-457.
16. OPH was cloned, expressed, purified, and stored at -80°C. It was thawed at 4°C before use and dialysed in pH 7.2, 0.1 M HEPES. The activity of OPH was measured in 1 mM paraoxon in pH 9.0, 0.15 M CHES buffer at 25°C, in which one OPH unit is the active amount allowing 1.0 μmol paraoxon to be hydrolyzed per minute. Protein amounts were measured using Pierce BCA Assay Kit (Pierce, product no. 23227). OPH stock solution, which was diluted 20-50 times by pH 7.5, 20 mM HEPES for activity measurements, had an initial specific activity of 1928 units/mg of protein.
17. -1 on an Eppendorf Thermomixer 5436 at 25°C for 2 h, and then centrifuging at 12000 rpm for 10 min. Repeatedly, the resulting OPH-FMS deposit was shaken vigorously and washed in pH 7.5, 20 mM HEPES (≥ 10 × 1.0 mL) at room temperature (21 ± 1°C and centrifuged to remove any nonfirmly immobilized enzyme, which was monitored in later cycles until both the protein and activity were undetectable in the supernatant. Finally, the OPH-FMS composite was thoroughly resuspended in the same washing buffer with 100 μL of the buffer per mg of FMS for further protein and activity measurement. To avoid any scattering during spectrophotometric measurement of protein amounts, a centrifugation step was taken to remove the silica particles after incubation of the OPH-FMS composite with the working reagent. The errors for both the protein amount and activity measurements were less than 5%.
18. (a) Zhou, H.-X.; Dill, K. A. Biochemistry 2001, 40, 11289-11293.
19. (b) Minton, A. P. J. Biol. Chem. 2001, 276, 10577-10580 and references therein.