ISSN:
1432-0614
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
,
Process Engineering, Biotechnology, Nutrition Technology
Notes:
Abstract The crystalline cell surface layer (S-layer) from Bacillus stearothermophilis PV72 was used as a matrix for reversible immobilization of β-d-galactosidase via disulphide bonds. In order to obtain an immobilization matrix stable towards acid, alkali and reducing agents such as dithiothreitol (DTT), the S-layer subunits were first cross-linked with glutaraldehyde. This was done in a way whereby 75% of the free amino groups remained unmodified, and then could be completely converted into sulphhydryl groups upon reaction with the monofunctional imidoester iminothiolane. After activation of the sulphhydryl groups with 2,2′-dipyridyldisulphide, 550 μg β-d-galactosidase could be immobilized per milligram of S-layer protein, which corresponds to one β-d-galactosidase molecule [relative molecular mass (Mr), 116000] per two S-layer subunits (Mr, 130 000). At least 90% of the sulphhydryl groups from the S-layer protein could be regenerated for further activation by cleaving the disulphide bonds with DTT. In comparative studies β-d-galactosidase was linked to carbodiimide-activated carboxyl groups of the S-layer protein.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00174459
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