Preparation of Active and Stable Biocatalytic Hydrogels for Use in Selective Transformations

Chemistry of Materials

Volumn 10, Issue 4, 1998, Pages 955-958

  Novick, Scott J ^a   Dordick, Jonathan S ^a,b  

^a IA United States   (United States)

^b UNIVERSITY OF IOWA   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

EID: 0000664454     PISSN: 08974756     EISSN: None     Source Type: Journal    
DOI: 10.1021/cm9708123     Document Type: Article

References (24)

1. Hartmeier, W. Immobilized biocatalysts: An introduction; Springer-Verlag: New York, 1988; pp 82-98.
2. Guilbault, G. Analytical uses of immobilized enzymes; Marcel Dekkar, Inc.: New York, 1984.
3. Gombotz, W. R.; Hoffman, A. S. Immobilization of biomolecules and cells on and within synthetic polymer hydrogels; Gombotz, W. R., Hoffman, A. S., Eds.; CRC Press: Boca Raton, FL, 1986; pp 95-126.
4. Taylor, R. F. Protein immobilization: Fundamentals and applications, 1st ed.; Marcel Dekker: New York, 1991; pp 93-98.
5. Taylor, R. F. Protein immobilization: Fundamentals and applications, 1st ed.; Marcel Dekker: New York, 1991; pp 15-29.
6. Dordick, J. S.; Linhardt, R. J.; Rethwisch, D. G. Chemtech 1994, 24, 33.
7. Mosbach, K. Immobilized enzymes. In Methods of enzymology; Colowick, S. P., Kaplan, N. O., Eds.; 1976, 44, 19-53.
8. Trevan, M. D. Immobilized Enzymes; John Wiley & Sons Ltd.: New York, 1980; pp 2-7.
9. Zaborsky, O. Immobilized Enzymes; CRC Press: Cleveland, OH, 1973; pp 5-48.
10. Rosevear, A.; Kennedy, J. F.; Cabral, J. M. S. Immobilised enzymes and cells; Adam Hilger: Bristol, 1987; pp 114-119.
11. Patil, N. S.; Dordick, J. S.; Rethwisch, D. G. Biomaterials 1996, 17, 2343.
12. Martinek, K.; Klibanov, A. M.; Goldmacher, V. S.; Berezin, I. V. Biochim. Biophys. Acta 1977, 485, 1.
13. After modification with acryloyl chloride, unreacted reagents were removed from the enzyme solution via ultrafiltration (Amicon YM-10 filter, 10 000 MWCO, running buffer consisted of 50 mM, pH 7,8 Tris buffer), and the enzyme was then lyophilized. The activity of the modified enzyme was measured in 50 mM pH 7.8 Tris buffer via the hydrolysis of the chromogenic synthetic tetrapeptide, N-succ-AAPF-p-nitroanilide. The initial rate of formation of free p-nitroaniline was measured at 410 nm. Modified CT retained at least 80% of the native enzyme activity.
14. Sucrose acrylate monomer (940 mg), cross-linker (5 mg), and the modified enzyme (1 mg) were dissolved in 5 mL of 50 mM pH 7.8 Tris buffer. The redox initiators, sodium persulfate and TEMED, were then added at a concentration of 1 wt % each. Polymerization was initiated by aspirating the solution to remove oxygen. Polymerization was carried out over an ice bath to reduce potential inactivation of the enzyme. For CT and SC, N-glutaryl-L-phe-p-nitroanilide was added as a substrate during free radical polymerization to protect the active site from inactivation and to reduce potential autolysis during the process.
15. 12
16. 12
17. I of 1.0.
18. Sugars are known to protect enzymes during the lyophilization process (Dabulis, K.; Klibanov, A. M. Biotechnol. Bioeng. 1993, 41, 566 ). Sugars have also been shown to stabilize enzymes in aqueous environments (Gray, C. J. Biocatalysis 1988, 187, 1).
19. Sugars are known to protect enzymes during the lyophilization process (Dabulis, K.; Klibanov, A. M. Biotechnol. Bioeng. 1993, 41, 566 ). Sugars have also been shown to stabilize enzymes in aqueous environments (Gray, C. J. Biocatalysis 1988, 187, 1).
20. Ugarova, N. N.; Rozhkova, G. D.; Berezin, I. V. Biochim. Biophys. Acta 1979, 570, 31.
21. o).
22. Zaks, A.; Klibanov, A. M. J. Biol. Chem. 1988, 263, 8017.
23. Gabel, D. FEBS Lett. 1974, 49, 280.
24. Kawamura, Y.; Nakanishi, K.; Matsuno, R.; Kamikubo, T. Biotechnol. Bioeng. 1981, 23, 1219.