title: Enzymatic activity of immobilized yeast phosphoglycerate kinase.
abstract: This work reports the first evidence that recombinant yeast phosphoglycerate kinase (PGK) is still significantly active when immobilized on glass and muscovite mica. Using previous work to improve the sensitivity of the existing setup, Tapping Mode atomic force microscopy (AFM) was used in a liquid environment to determine the surface enzyme coverage of derivatized mica and glass slides. When associated to spectrophotometric measurements, the AFM data allows assessing the catalytic constant of surface enzymes and comparing it to bulk values. The validity of the Michaelis-Menten model for surface reactions is discussed, supported by spectroscopic measurements of the surface consumption of 1,3-bis-phosphoglycerate (1,3-BPG). Only a few percent of the enzyme material maintains its initial bulk activity. This value could constitute a guideline for biosensors made with the method used here whenever a rapid assessment of the remaining surface activity is needed.
authors.affiliation: Centre de Physique Moléculaire Optique et Hertzienne, 33600 Pessac, France. churth@mail.utexas.edu <churth@mail.utexas.edu>
authors.forename: Cedric
authors.initials: C
authors.lastname: Hurth
authors.forename: Chantal
authors.initials: C
authors.lastname: Tassius
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Talbot
authors.forename: Abdelhamid
authors.initials: A
authors.lastname: Maali
authors.forename: Cendrine
authors.initials: C
authors.lastname: Moskalenko
authors.forename: Philippe
authors.initials: P
authors.lastname: Minard
authors.forename: Jean-Pierre
authors.initials: JP
authors.lastname: Aimé
authors.forename: Françoise
authors.initials: F
authors.lastname: Argoul
mesh_terms: D000538:Aluminum Silicates / Q000737:chemistry; D001672:Biocompatible Materials / Q000737:chemistry*; D015374:Biosensing Techniques / Q000379:methods*; D004789:Enzyme Activation; D004800:Enzymes, Immobilized / Q000737:chemistry; D005898:Glass / Q000737:chemistry; D010735:Phosphoglycerate Kinase / Q000737:chemistry* / Q000648:ultrastructure; D013499:Surface Properties; D015003:Yeasts / Q000201:enzymology*
chemical_list: D000538:Aluminum Silicates; D001672:Biocompatible Materials; D004800:Enzymes, Immobilized; D010735:Phosphoglycerate Kinase; C011934:mica