Biopolym. Cell. 2015; 31(4):301-308.
Molecular and Cell Biotechnologies
SPR investigations of the formation of intermediate layer of the immunosensor bioselective element based on the recombinant Staphylococcal protein A
1Rachkov A. E., 1, 2Bakhmachuk A. O., 1Gorbatiuk O. B., 1, 3Matsishin M. J., 4Khristosenko R. V., 4Ushenin Iu. V., 1, 3Soldatkin A. P.
  1. Institute of Molecular Biology and Genetics, NAS of Ukraine,
    150, Akademika Zabolotnoho Str., Kyiv, Ukraine, 03680
  2. Educational and Scientific Center "Institute of Biology",
    Taras Shevchenko National University of Kyiv
    64/13, Volodymyrska Str., Kyiv, Ukraine, 01601
  3. Institute of High Technologies,
    Taras Shevchenko National University of Kyiv
    2, korp.5, Pr. Akademika Hlushkova, Kyiv, Ukraine, 03022
  4. V. Ye. Lashkaryov Institute of Semiconductor Physics, NAS of Ukraine
    41, Prospect Nauki, Kyiv, Ukraine, 03028

Abstract

Aim. To investigate the formation of an intermediate layer of the immunosensor bioselective element based on the recombinant protein A from Staphylococcus aureus with cysteine residue (SPA-Cys) and its interactions with human IgG using the SPR spectrometer «Plasmon». Methods. The activity of the immune components applied was tested by ELISA. The spectrometry of surface plasmon resonance was used for studying protein immobilization on a gold sensor surface and interactions between the immobilized SPA-Cys and human immunoglobulin. Results. A direct dependence of the sensor response on the concentration of SPA-Cys in the range of 0.2 to 2 µM at its immobilization was demonstrated. The efficiency of blocking nonspecific adsorption sites on the sensor surface with milk proteins and the direct dependence of the sensor response on IgG concentration and surface density of immobilized SPA-Cys were shown. Fitting the experimental data to a Langmuir plot yields a Kd value for SPA-Cys/IgG binding 8.5 ± 0.7× 10-8 M (Ka = 1.2 ± 0.1× 107 M–1). The determined equilibrium binding constant indicates a quite strong interaction and its value is consistent with the literature data. Conclusions. A successful immobilization of SPA-Cys on a gold surface of the SPR spectrometer while preserving its high immunoglobulin-binding activity, selectivity and stability of the sensor response confirms the efficiency of SPA-Cys as an intermediate component for the creation of the immunosensor bioselective elements.
Keywords: immunoglobulin, recombinant Staphylococcal protein A, surface plasmon resonance, protein immobilization, immunosensor, equilibrium binding constant.

References

[1] Soldatkin AP, Dzyadevych SV, Korpan YI, Sergeyeva TA, Arkhypova VN, Biloivan OA, Soldatkin OO, Shkotova LV, Zinchenko OA, Peshkova VM, Saiapina OY, Marchenko SV, El'skaya AV. Biosensors. A quarter of a century of R&D experience. Biopolym Cell. 2013; 29(3):188–206.
[2] Ramanaviciene A, German N, Kausaite-Minkstimiene A, Voronovic J, Kirlyte J, Ramanavicius A. Comparative study of surface plasmon resonance, electrochemical and electroassisted chemiluminescence methods based immunosensor for the determination of antibodies against human growth hormone. Biosens Bioelectron. 2012;36(1):48-55.
[3] de Juan-Franco E, Caruz A, Pedrajas JR, Lechuga LM. Site-directed antibody immobilization using a protein A-gold binding domain fusion protein for enhanced SPR immunosensing. Analyst. 2013;138(7):2023-31.
[4] Makaraviciute A, Ramanaviciene A. Site-directed antibody immobilization techniques for immunosensors. Biosens Bioelectron. 2013;50:460-71.
[5] Wang C, Feng B. [Research progress on site-oriented and three-dimensional immobilization of proteins]. Mol Biol (Mosk). 2015;49(1):3-25.
[6] Abrahmsén L, Moks T, Nilsson B, Hellman U, Uhlén M. Analysis of signals for secretion in the staphylococcal protein A gene. EMBO J. 1985;4(13B):3901-6.
[7] Moks T, Abrahmsén L, Nilsson B, Hellman U, Sjöquist J, Uhlén M. Staphylococcal protein A consists of five IgG-binding domains. Eur J Biochem. 1986;156(3):637-43.
[8] Sjodahl J. Repetitive sequences in protein A from Staphylococcus aureus. Arrangement of five regions within the protein, four being highly homologous and Fc-binding. Eur J Biochem. 1977;73(2):343-51.
[9] Sidorin EV, Solov'eva TF. IgG-binding proteins of bacteria. Biochemistry (Mosc). 2011;76(3):295-308.
[10] Muramatsu H, Dicks JM, Tamiya E, Karube I. Piezoelectric crystal biosensor modified with protein A for determination of immunoglobulins. Anal Chem. 1987;59(23):2760-3.
[11] Prusak-Sochaczewski E, Luong JHT. A new approach to the development of a reusable piezoelectric crystal biosensor. Anal Lett. 1990; 23(3):401–9.
[12] Kanno S, Yanagida Y, Haruyama T, Kobatake E, Aizawa M. Assembling of engineered IgG-binding protein on gold surface for highly oriented antibody immobilization. J Biotechnol. 2000;76(2-3):207-14.
[13] Ljungquist C, Jansson B, Moks T, Uhlén M. Thiol-directed immobilization of recombinant IgG-binding receptors. Eur J Biochem. 1989;186(3):557-61.
[14] Gorbatiuk OB, Bakhmachuk AO, Dubey LV, Usenko MO, Irodov DM, Okunev OV, Kostenko OM, Rachkov AE, Kordium VA. Recombinant Staphylococcal protein A with cysteine residue for preparation of affinity chromatography stationary phase and immunosensor applications. Biopolym Cell. 2015; 31(2):115–22.
[15] Homola J. Surface plasmon resonance sensors for detection of chemical and biological species. Chem Rev. 2008;108(2):462-93.
[16] Rengevich OV, Shirshov YuM, Ushenin YuV, Beketov AG. Separate determination of thickness and optical parameters by surface plasmon resonance: accuracy consideration. Semiconductor physics, quantum electronics and optoelectronics. 1999; 2(2):28–35.
[17] Gridina N, Dorozinsky G, Khristosenko R, Maslov V, Samoylov A, Ushenin Yu, Shirshov Yu. Surface plasmon resonance biosensor. Sensors & Transducers Journal. 2013; 149(2): 60–8.
[18] Gorbatiuk OB, Okunev OV, Nikolaev YuS, Svyatenko OV, Kordium VA. Construction, expression, functional characterization and practical application of fusion protein SPA-BAPmut. Biopolym Cell. 2013; 29(1):49–54.
[19] Rachkov A, Holodova Y, Ushenin Y, Miroshnichenko D, Telegeev G, Soldatkin A. Development of bioselective element of SPR spectrometer for monitoring of oligonucleotide interactions and comparison with thermodynamic calculations. Sens Lett. 2009;7(5):957–61.
[20] Karlsson R, Ståhlberg R. Surface plasmon resonance detection and multispot sensing for direct monitoring of interactions involving low-molecular-weight analytes and for determination of low affinities. Anal Biochem. 1995;228(2):274-80.
[21] Goode JA, Rushworth JV, Millner PA. Biosensor Regeneration: A Review of Common Techniques and Outcomes. Langmuir. 2015;31(23):6267-76.
[22] Sergeyeva TA, Soldatkin AP, Rachkov AE, Tereschenko MI, Piletsky SA, El`skaya AV. β-Lactamase label-based potentiometric biosensor for α-2 interferon detection. Anal Chim Acta. 1999;390(1-3):73–81.
[23] Schwartz MP, Alvarez SD, Sailor MJ. Porous SiO2 interferometric biosensor for quantitative determination of protein interactions: binding of protein A to immunoglobulins derived from different species. Anal Chem. 2007;79(1):327-34.
[24] Lindmark R, Biriell C, Sjöquist J. Quantitation of specific IgG antibodies in rabbits by a solid-phase radioimmunoassay with 125I-protein A from Staphylococcus aureus. Scand J Immunol. 1981;14(4):409-20.
[25] Saha K, Bender F, Gizeli E. Comparative study of IgG binding to proteins G and A: nonequilibrium kinetic and binding constant determination with the acoustic waveguide device. Anal Chem. 2003;75(4):835-42.