Polyethylene glycol as spacer for solid phase enzyme immobilization

ENZIMAS

INIMOVILIZACION DE ENZIMAS

GLICOL DE POLIETILENO

BIBLIOGRAFIA NACIONAL QUIMICA

With the aim to improve the performance of enzyme bound to hydrophilic solid phases, their immobilization with polyethylene glycol (PEG) tether have been studied. Sweet potato
-amylase, which hydrolyses the high molecular weight substrate starch and
-galactosidase, which acts on low molecular weight substrates, were used as model enzymes and beaded thiol–agarose as solid phase. Several two step methods for the introduction of the tether using a bis-oxirane homobifunctional PEG as well as a heterobifunctional derivative with a hydroxysuccinimide ester and a maleimide group have been evaluated. Amino groups, native and de novo thiol groups in the enzymes were utilized for immobilization. The best approach was found to be to first introduce the PEG derivative via one of its reactive groups to the enzyme. Subsequently the formed conjugate was bound to the solid phase by the remaining reactive group. Attempts to first introduce the PEG tether into the solid phase were not successful. Ahigh degree of substitution with PEG chains on the enzyme leads to high immobilization yields for both
-amylase and
-galactosidase, but relatively lower gel-bound activity for the former enzyme which is acting on a high molecular weight substrate and thus more sensitive for steric shielding effects. With optimal degree of PEG substitution (which occurred at five times molar excess of the heterobifunctional reagent) the gel-bound activity of
-amylase was increased from 12% (for the derivative without tether) to 31%.

Enzyme and Microbial Technology v. 33, no. 7, 2003. -- p. 890-898

Elsevier

2003

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DOI:10.1016/S0141-0229(03)00221-7