Hydrolysis of a Lipid Membrane by Single Enzyme Molecules: Accurate Determination of Kinetic Parameters.
Artikel i vetenskaplig tidskrift, 2015

The accurate determination of the maximum turnover number and Michaelis constant for membrane enzymes remains challenging. Here, this problem has been solved by observing in parallel the hydrolysis of thousands of individual fluorescently labeled immobilized liposomes each processed by a single phospholipase A2 molecule. The release of the reaction product was tracked using total internal reflection fluorescence microscopy. A statistical analysis of the hydrolysis kinetics was shown to provide the Michaelis-Menten parameters with an accuracy better than 20 % without variation of the initial substrate concentration. The combined single-liposome and single-enzyme mode of operation made it also possible to unravel a significant nanoscale dependence of these parameters on membrane curvature.

membranes

phospholipases

liposomes

kinetics

enzyme catalysis

Författare

Michael Rabe

Chalmers, Teknisk fysik, Biologisk fysik

Seyed Tabaei

Chalmers, Teknisk fysik, Biologisk fysik

Henrik Zetterberg

Göteborgs universitet

Vladimir Zhdanov

Russian Academy of Sciences

Fredrik Höök

Chalmers, Teknisk fysik, Kemisk fysik

Angewandte Chemie - International Edition

1433-7851 (ISSN) 1521-3773 (eISSN)

Vol. 54 1022-1026

Ämneskategorier

Neurovetenskaper

DOI

10.1002/anie.201409603

PubMed

25429738