By the same authors

From the same journal

From the same journal

Comparative Assessment of Different Histidine-Tags for Immobilization of Protein onto Surface Plasmon Resonance Sensorchips

Research output: Contribution to journalArticle

Standard

Comparative Assessment of Different Histidine-Tags for Immobilization of Protein onto Surface Plasmon Resonance Sensorchips. / Fischer, Marcus; Leech, Andrew P.; Hubbard, Roderick E.

In: Analytical Chemistry, Vol. 83, No. 5, 01.03.2011, p. 1800-1807.

Research output: Contribution to journalArticle

Harvard

Fischer, M, Leech, AP & Hubbard, RE 2011, 'Comparative Assessment of Different Histidine-Tags for Immobilization of Protein onto Surface Plasmon Resonance Sensorchips', Analytical Chemistry, vol. 83, no. 5, pp. 1800-1807. https://doi.org/10.1021/ac103168q

APA

Fischer, M., Leech, A. P., & Hubbard, R. E. (2011). Comparative Assessment of Different Histidine-Tags for Immobilization of Protein onto Surface Plasmon Resonance Sensorchips. Analytical Chemistry, 83(5), 1800-1807. https://doi.org/10.1021/ac103168q

Vancouver

Fischer M, Leech AP, Hubbard RE. Comparative Assessment of Different Histidine-Tags for Immobilization of Protein onto Surface Plasmon Resonance Sensorchips. Analytical Chemistry. 2011 Mar 1;83(5):1800-1807. https://doi.org/10.1021/ac103168q

Author

Fischer, Marcus ; Leech, Andrew P. ; Hubbard, Roderick E. / Comparative Assessment of Different Histidine-Tags for Immobilization of Protein onto Surface Plasmon Resonance Sensorchips. In: Analytical Chemistry. 2011 ; Vol. 83, No. 5. pp. 1800-1807.

Bibtex - Download

@article{58c307c07634428a9d6534775350af14,
title = "Comparative Assessment of Different Histidine-Tags for Immobilization of Protein onto Surface Plasmon Resonance Sensorchips",
abstract = "Surface plasmon resonance (SPR) is widely used to assess the kinetics and thermodynamics of binding of two molecules. The major challenge is immobilization of one molecule onto the sensorchip for robust detection of binding of the other molecule. We have compared a number of immobilization strategies for noncovalent attachment of an example protein (the substrate binding protein SiaP) by hexa-histidine (His), deca-His, and double-His tags to a nickel-nitrilotriacetic acid (NTA) surface. The stability of immobilization was assessed, and the binding of two low molecular weight ligands, Neu5Ac and 2-keto-3-deoxy-D-glycero-D-galacto-nononic acid (KDN), at different temperatures studied. The hexa-His tagged SiaP washed off from the surface too rapidly for ligand binding to be measured reliably. Systematic variation of chip loading identified conditions under which the deca-His tagged SiaP could generate reliable results. The double-His tagged protein performed as well as covalently attached deca-His tagged protein at 15, 25, and 35 C. The observed ligand binding kinetics were comparable for all immobilization strategies, and thermodynamic values calculated from SPR are in agreement with solution-based isothermal titration calorimetry measurements. Extended trials suggest that covalent attachment is preferable for screening campaigns, whereas the double-His-tag strategy allows rapid regeneration of the chip, for example, when tight binding compounds are assessed.",
keywords = "HAEMOPHILUS-INFLUENZAE, BINDING, TRANSPORTER, INHIBITORS, DISCOVERY, SIAP",
author = "Marcus Fischer and Leech, {Andrew P.} and Hubbard, {Roderick E.}",
year = "2011",
month = "3",
day = "1",
doi = "10.1021/ac103168q",
language = "English",
volume = "83",
pages = "1800--1807",
journal = "Analytical Chemistry",
issn = "0003-2700",
publisher = "American Chemical Society",
number = "5",

}

RIS (suitable for import to EndNote) - Download

TY - JOUR

T1 - Comparative Assessment of Different Histidine-Tags for Immobilization of Protein onto Surface Plasmon Resonance Sensorchips

AU - Fischer, Marcus

AU - Leech, Andrew P.

AU - Hubbard, Roderick E.

PY - 2011/3/1

Y1 - 2011/3/1

N2 - Surface plasmon resonance (SPR) is widely used to assess the kinetics and thermodynamics of binding of two molecules. The major challenge is immobilization of one molecule onto the sensorchip for robust detection of binding of the other molecule. We have compared a number of immobilization strategies for noncovalent attachment of an example protein (the substrate binding protein SiaP) by hexa-histidine (His), deca-His, and double-His tags to a nickel-nitrilotriacetic acid (NTA) surface. The stability of immobilization was assessed, and the binding of two low molecular weight ligands, Neu5Ac and 2-keto-3-deoxy-D-glycero-D-galacto-nononic acid (KDN), at different temperatures studied. The hexa-His tagged SiaP washed off from the surface too rapidly for ligand binding to be measured reliably. Systematic variation of chip loading identified conditions under which the deca-His tagged SiaP could generate reliable results. The double-His tagged protein performed as well as covalently attached deca-His tagged protein at 15, 25, and 35 C. The observed ligand binding kinetics were comparable for all immobilization strategies, and thermodynamic values calculated from SPR are in agreement with solution-based isothermal titration calorimetry measurements. Extended trials suggest that covalent attachment is preferable for screening campaigns, whereas the double-His-tag strategy allows rapid regeneration of the chip, for example, when tight binding compounds are assessed.

AB - Surface plasmon resonance (SPR) is widely used to assess the kinetics and thermodynamics of binding of two molecules. The major challenge is immobilization of one molecule onto the sensorchip for robust detection of binding of the other molecule. We have compared a number of immobilization strategies for noncovalent attachment of an example protein (the substrate binding protein SiaP) by hexa-histidine (His), deca-His, and double-His tags to a nickel-nitrilotriacetic acid (NTA) surface. The stability of immobilization was assessed, and the binding of two low molecular weight ligands, Neu5Ac and 2-keto-3-deoxy-D-glycero-D-galacto-nononic acid (KDN), at different temperatures studied. The hexa-His tagged SiaP washed off from the surface too rapidly for ligand binding to be measured reliably. Systematic variation of chip loading identified conditions under which the deca-His tagged SiaP could generate reliable results. The double-His tagged protein performed as well as covalently attached deca-His tagged protein at 15, 25, and 35 C. The observed ligand binding kinetics were comparable for all immobilization strategies, and thermodynamic values calculated from SPR are in agreement with solution-based isothermal titration calorimetry measurements. Extended trials suggest that covalent attachment is preferable for screening campaigns, whereas the double-His-tag strategy allows rapid regeneration of the chip, for example, when tight binding compounds are assessed.

KW - HAEMOPHILUS-INFLUENZAE

KW - BINDING

KW - TRANSPORTER

KW - INHIBITORS

KW - DISCOVERY

KW - SIAP

UR - http://www.scopus.com/inward/record.url?scp=79952174208&partnerID=8YFLogxK

U2 - 10.1021/ac103168q

DO - 10.1021/ac103168q

M3 - Article

VL - 83

SP - 1800

EP - 1807

JO - Analytical Chemistry

JF - Analytical Chemistry

SN - 0003-2700

IS - 5

ER -