A piezoelectric quartz crystal biosensor: the use of two single cysteine mutants of the periplasmic Escherichia coli glucose/galactose receptor as target proteins for the detection of glucose.
We have examined the potential utility of a glucose biosensor that employs the glucose/galactose receptor of Escherichia coli with a quartz crystal microbalance (QCM). Two different genetically engineered mutant proteins were utilized, each involving the incorporation of a single cysteine into the amino acid sequence of the protein. The proteins were immobilized on the surface of a piezoelectric crystal by a direct sulfur-gold linkage. Since the cysteines were located at different positions in the sequence, the receptors attach to the surface with different orientations. Considering only mass effects, the target sugars for this receptor are predicted to be too small to be detectable with a QCM. However, our sensors indicated measurable and reproducible frequency responses when immobilized receptor was exposed to sugar. This unexpectedly large frequency response occurs because the protein film is transformed from a viscous layer to a more rigid nondissipative film. The QCM can detect these changes because of the direct linkage of the proteins to the surface. Calculations of the frequency response expected for a viscoelastic film with different rheological characteristics support this hypothesis. This study is significant because it illustrates a widened applicability for the QCM methodology to protein systems that bind small molecules and undergo ligand-induced conformational changes.
Carmon
Kendra S
KS
Department of Chemistry, Clarkson University, Potsdam, New York 13699, USA.
Baltus
Ruth E
RE
Luck
Linda A
LA
D000409:Alanine / Q000235:genetics; D019943:Amino Acid Substitution / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D003460:Crystallization; D003545:Cysteine / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D005947:Glucose / Q000378:metabolism*; D008024:Ligands; D016297:Mutagenesis, Site-Directed; D019897:Periplasm / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics; D011487:Protein Conformation; D011791:Quartz; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000378:metabolism; D013499:Surface Properties
D029968:Escherichia coli Proteins; D008024:Ligands; D011956:Receptors, Cell Surface; C039707:galactose receptor; C027786:glucose receptor; D011791:Quartz; D005947:Glucose; D003545:Cysteine; D000409:Alanine