title: Characterization of the specific interaction between the DNA aptamer sgc8c and protein tyrosine kinase-7 receptors at the surface of T-cells by biosensing AFM.
abstract: We studied the interaction of the specific DNA aptamer sgc8c immobilized at the AFM tip with its corresponding receptor, the protein tyrosine kinase-7 (PTK7) embedded in the membrane of acute lymphoblastic leukemia (ALL) cells (Jurkat T-cells). Performing single molecule force spectroscopy (SMFS) experiments, we showed that the aptamer sgc8c bound with high probability (38.3 ± 7.48%) and high specificity to PTK7, as demonstrated by receptor blocking experiments and through comparison with the binding behavior of a nonspecific aptamer. The determined kinetic off-rate (koff= 5.16 s-1) indicates low dissociation of the sgc8c-PTK7 complex. In addition to the pulling force experiments, simultaneous topography and recognition imaging (TREC) experiments using AFM tips functionalized with sgc8c aptamers were realized on the outer regions surface of surface-immobilized Jurkat cells for the first time. This allowed determination of the distribution of PTK7 without any labeling and at near physiological conditions. As a result, we could show a homogeneous distribution of PTK7 molecules on the outer regions of ALL cells with a surface density of 325 ± 12 PTK7 receptors (or small receptor clusters) per μm2. Graphical Abstract The specific interaction of the DNA aptamer sgc8c and protein tyrosine kinase-7 (PTK7) on acute lymphoblastic leukemia (ALL) cells was characterized. AFM based single molecule force spectroscopy (SMFS) yielded a kinetic off-rate of 5.16 s-1of the complex. Simultaneous topography and recognition imaging (TREC) revealed a PTK7 density of 325 ± 12 molecules or clusters per μm2in the cell membrane.
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020, Linz, Austria.
authors.forename: Michael
authors.initials: M
authors.lastname: Leitner
authors.affiliation: Faculty of Mathematics, Physics, and Informatics, Comenius University, Mlynska dolina F1, 842 48, Bratislava, Slovakia.|Institute of Biochemistry and Animal Genetics, Slovak Academy of Sciences, Moyzesova 61, 900 28, Ivanka pri Dunaji, Slovakia.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Poturnayova
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020, Linz, Austria.
authors.forename: Constanze
authors.initials: C
authors.lastname: Lamprecht
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020, Linz, Austria.
authors.forename: Sabine
authors.initials: S
authors.lastname: Weich
authors.affiliation: Institute of Biochemistry and Animal Genetics, Slovak Academy of Sciences, Moyzesova 61, 900 28, Ivanka pri Dunaji, Slovakia.
authors.forename: Maja
authors.initials: M
authors.lastname: Snejdarkova
authors.affiliation: Faculty of Mathematics, Physics, and Informatics, Comenius University, Mlynska dolina F1, 842 48, Bratislava, Slovakia.
authors.forename: Ivana
authors.initials: I
authors.lastname: Karpisova
authors.affiliation: Faculty of Mathematics, Physics, and Informatics, Comenius University, Mlynska dolina F1, 842 48, Bratislava, Slovakia.
authors.forename: Tibor
authors.initials: T
authors.lastname: Hianik
authors.affiliation: Institute of Biophysics, Johannes Kepler University Linz, Gruberstrasse 40, 4020, Linz, Austria. andreas.ebner@jku.at.
authors.forename: Andreas
authors.initials: A
authors.lastname: Ebner
mesh_terms: D052157:Aptamers, Nucleotide / Q000378:metabolism*; D001665:Binding Sites; D015374:Biosensing Techniques / Q000379:methods; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D006801:Humans; D019169:Jurkat Cells; D018625:Microscopy, Atomic Force / Q000379:methods*; D057054:Molecular Imaging / Q000379:methods*; D011485:Protein Binding; D025941:Protein Interaction Mapping / Q000379:methods*; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D013601:T-Lymphocytes / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D052157:Aptamers, Nucleotide; D015815:Cell Adhesion Molecules; C096905:PTK7 protein, human; D020794:Receptor Protein-Tyrosine Kinases
keywords: DNA aptamer; Energy landscape; Molecular recognition; PTK7; Recognition imaging; Single molecule force spectroscopy; T-cell