Escherichia coli TEM1 beta-lactamase in CTAB reverse micelles: exchange/diffusion-limited catalysis.
We report kinetic data of penicillin hydrolysis catalyzed by beta-lactamase entrapped in reverse micelles formed with cetyl trimethylammonium bromide (CTAB), n-octane, hexanol and aqueous buffer. The K(cat) of this diffusion-limited reaction can be improved in aqueous buffer by a factor of 1.1-1.2 just by increasing the phosphate buffer concentration from 50 to 100 mM. In reverse micelles, increasing the buffer concentration has little effect on K(cat) when the size of the empty micelle is below the size of the protein. However, in larger micelles, the effect is enhanced and the K(cat) improves several fold, changing the form of the curve of K(cat) versus Wo from bell-shaped to almost hyperbolic. The results indicate that micellar exchange and internal diffusion may limit the reaction in reverse micelles and provide further evidence that the form of the curve depends on other factors besides the relationship between the size of the enzyme and that of the empty reverse micelle.
Moreno-Hagelsieb
G
G
Dpto. de Reconocimiento Molecular y Bioestructura, Instituto de Biotecnología-UNAM, Av. Universidad 2001, Cuernavaca, Morelos, Mexico. moreno@ibt.unam.mx
Gómez-Puyou
A
A
Soberon
X
X
D002384:Catalysis; D000077286:Cetrimonium; D002593:Cetrimonium Compounds / Q000378:metabolism*; D004058:Diffusion; D004926:Escherichia coli / Q000201:enzymology*; D008823:Micelles; D010406:Penicillins / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism; D017680:Potassium Compounds / Q000378:metabolism; D001618:beta-Lactamases / Q000378:metabolism*
D002593:Cetrimonium Compounds; D008823:Micelles; D010406:Penicillins; D010710:Phosphates; D017680:Potassium Compounds; C013216:potassium phosphate; D001618:beta-Lactamases; C034413:beta-lactamase TEM-1; D000077286:Cetrimonium