PRODUCTION OF MYCOBACTERIUM TUBERCULOSIS TB10.4 RECOMBINANT PROTEIN IN ESCHERICHIA COLI

Cover Page

Cite item

Abstract

Nowadays tuberculosis is considered one of the most dangerous infectious diseases occurring everywhere, and it remains a cause of death of millions of people around the world. According to the World Health Organization data, in 2013 tuberculosis caused more than 9 million cases worldwide and about 1.5 million of infected people died. The causative agent of tuberculosis in most cases is Mycobacterium tuberculosis. But sometimes it can be Mycobacterium bovis or Mycobacterium africanum. Mainly as a result of infection, a bacterial infection affects the lungs, but the disease may develop in other organs and tissues. Now for the prevention of tuberculosis vaccination of newborns with attenuated vaccine BCG is widely used. The production of this vaccine is cheap and it is safe to use. Thus today, vaccination is the primary means of prevention of tuberculosis. However dubious efficacy and a number of side effects observed after vaccination, makes the scientific community to develop new effective methods for the treatment of tuberculosis. One of the ways to develop new vaccines against tuberculosis is to provide a subunit vaccine based on recombinant proteins. Advantages of subunit vaccines are that the preparation containing the purified protein is stable and secure, its chemical properties are known, it does not contain additional proteins and nucleic acids, which could cause undesirable effects in the human body. One of the most promising antigens for use as components in new vaccines is considered a low molecular weight secreted protein TB10.4. TB10.4 protein is recognized at an early stage of tuberculous infection and contributes to the proliferation of lymphocytes responsible for the production of IFNγ. TB10.4 protein also possesses an adjuvant effect when administered in combination with mycobacterial proteins. Given these properties, the recombinant protein TB10.4 can be used to generate new candidate vaccines against tuberculosis. During the study was created high-yield E. coli strain, which produces the recombinant protein TB10.4, selected the optimal protocol of induction of the gene encoding the protein. The protein was purified using metal affinity chromatography. The purity of the final preparation reached 98%. 

About the authors

I. V. Dukhovlinov

Research Institute of Highly Pure Biopreparations, St. Petersburg, Russian Federation

Author for correspondence.
Email: dukhovlinov@gmail.com
Russian Federation

E. A. Fedorova

Research Institute of Highly Pure Biopreparations, St. Petersburg, Russian Federation

Email: fake@neicon.ru
Russian Federation

O. A. Dobrovolskaya

Research Institute of Highly Pure Biopreparations, St. Petersburg, Russian Federation

Email: fake@neicon.ru
Russian Federation

E. G. Bogomolova

Research Institute of Highly Pure Biopreparations, St. Petersburg, Russian Federation

Email: fake@neicon.ru
Russian Federation

E. N. Chernyaeva

Research Institute of Highly Pure Biopreparations, St. Petersburg, Russian Federation

Email: fake@neicon.ru
Russian Federation

R. I. Al-Shekhadat

Research Institute of Highly Pure Biopreparations, St. Petersburg, Russian Federation

Email: fake@neicon.ru
Russian Federation

A. S. Simbirtsev

Research Institute of Highly Pure Biopreparations, St. Petersburg, Russian Federation

Email: fake@neicon.ru
Russian Federation

References

  1. Поливалентные вакцины, содержащие рекомбинант ные вирусные векторы: пат. 012037 США: МПК C12N 7/01, C12N 15/34, C12N 15/31, C12N 15/861, A61K 39/04 / Хавенга М.Я.Э. (Нидерланды), Вогелс Р. (Нидерланды), Сэдофф Д. (США), Хоун Д. (США), Скейки Я.А.В. (США), Радошевич К. (Нидерланды); заявитель и патентообладатель Круселл Холланд Б.В. (Нидерланды), Эйрас Глоубал Тиби Вэксин Фаундейшн (США); патент. поверенный Медведев В.Н. (Россия). – № 200701084, заявл. 2005.11.15, опубл. 2006.05.26. [Polivalentnye vaktsiny, soderzhashchie rekombinantnye virusnye vektory [Polyvalent vaccines containing recombinant viral vectors]: pat. 012037 США: МПК C12N 7/01, C12N 15/34, C12N 15/31, C12N 15/861, A61K 39/04 / Khavenga M.Ya.E. (Netherlands), Vogels R. (Netherlands), Sedoff D. (USA), Khoun D. (USA), Skeiki Ya.A.V. (USA), Radoshevich K. (Netherlands); applicant and patentee Krusell Kholland B.V. (Netherlands), Eiras Global Tibi Vexin Foundation (USA); patent attorney Medvedev V.N. (Russia). – № 200701084, Appl. 2005.11.15, publ. 2006.05.26. (In Russ.)]
  2. Стукова M.A., Заболотных Н.В., Виноградова Т.И., Гергерт В.Я., Апт А.С., Капрельянц А.С., Ерохин В.В., Яблонский П.К., Киселев О.И. Профилактика туберкулеза: современные подходы к разработке противотуберкулезных вакцин // Вестник Российской академии медицинских наук. 2012. № 11. C. 45–52. [Stukova M.A., Zabolotnykh N.V., Vinogradova T.I., Gergert V.Ya., Apt A.S., Kaprel’yants A.S., Erokhin V.V., Yablonskii P.K., Kiselev O.I. Prevention of tuberculosis: current approaches to development of vaccines. Vestnik Rossiiskoi akademii meditsinskikh nauk = Herald of the Russian Academy of Medical Sciences, 2012, no. 11, pp. 45–52. (In Russ.)]
  3. Татьков С.И., Дейнеко Е.В., Фурман Д.П. Перспективы создания противотуберкулезных вакцин нового поколения // Вавиловский журнал генетики и селекции. 2011. Т. 15, № 1. С. 114–129. [Tat’kov S.I., Peineko E.V., Furman D.P. Prospects for designing a new generation of anti-tuberculosis vaccine. Vavilovskii zhurnal genetiki i selektsii = Vavilov Journal of Genetics and Breeding, 2011, vol. 15, no. 1, pp. 114–129. (In Russ.)]
  4. Cole S.T., Brosch R., Parkhill J., Garnier T., Churcher C., Harris D., Gordon S.V., Eiglmeier K., Gas S., Barry C.E., Tekaia F., Badcock K., Basham D., Brown D., Chillingworth T., Connor R., Davies R., Devlin K., Feltwell T., Gentles S., Hamlin N., Holroyd S., Hornsby T., Jagels K., Krogh A., McLean J., Moule S., Murphy L., Oliver K., Osborne J., Quail M.A., Rajandream M.A., Rogers J., Rutter S., Seeger K., Skelton J., Squares R., Squares S., Sulston J.E., Taylor K., Whitehead S., Barrell B.G. Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature, 1998, vol. 393, no. 6685, pp. 537–544.
  5. Haile M., Källenius G. Recent developments in tuberculosis vaccines. Curr. Opin. Infect. Dis., 2005, vol. 18, no. 3, pp. 211–215.
  6. Iem V., Somphavong S., Buisson Y., Steenkeste N., Breysse F., Chomarat M., Sylavanh P., Nanthavong P., Rajoharison A., Berland J.L., Paboriboune P. Resistance of Mycobacterium tuberculosis to antibiotics in Lao PDR: first multicentric study conducted in 3 hospitals. BMC Infect. Dis., 2013, vol. 13, no. 1, 275 p. doi: 10.1186/1471-2334-13-275
  7. Invitrogen. Ni-NTA purification system. User manual. Catalog nos. K950-01, K951-01, K952-01, K953-01, K954-01, R901-01, R901-10, R 901-15. Version C. 25-0496, 2006, 32 p.
  8. Lindenstrøm T., Agger E.M., Korsholm K.S., Darrah P.A., Aagaard C., Seder R.A., Rosenkrands I., Andersen P. Tuberculosis subunit vaccination provides long-term protective immunity characterized by multifunctional CD4 memory T cells. J. Immunol., 2009, vol. 182, no. 12, pp. 8047–8055. doi: 10.4049/jimmunol.0801592
  9. Majumder K. Ligation-free gene synthesis by PCR: synthesis and mutagenesis at multiple loci of a chimeric gene encoding OmpA signal peptide and hirudin. Gene, 1992, vol. 110, no. 1, pp. 89–94. doi: 10.1016/0378-1119(92)90448-X
  10. Rodrigues L.C., Pereira S.M., Cunha S.S., Genser B., Ichihara M.Y., de Brito S.C., Hijjar M.A., Dourado I., Cruz A.A., Sant’Anna C., Bierrenbach A.L., Barreto M.L. Effect of BCG revaccination on incidence of tuberculosis in school-aged children in Brazil: the BCG-REVAC cluster-randomised trial. Lancet, 2005, vol. 366, no. 9493, pp. 1290–1295. doi: 10.1016/S0140-6736(05)67145-0
  11. Russell D.G., Barry C.E., Flynn J.L. Tuberculosis: what we don’t know can, and does, hurt us. Science, 2010, vol. 328, no. 5980, pp. 852–856. doi: 10.1126/science.1184784
  12. Schägger H., von Jagow G. Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in the range from 1 to 100 kDa. Anal. Biochem., 1987, vol. 166, no. 2, pp. 368–379.
  13. Sharma A.K., Khuller G.K. Recombinant mycobacterial proteins future directions to improve protective efficacy. Indian J. Exp. Biol., 2001, vol. 39, no. 12, pp. 1214–1219.
  14. Skjøt R.L.V., Oettinger T., Rosenkrands I., Ravn P., Brock I., Jacobsen S., Andersen P. Comparative evaluation of low-molecularmass proteins from Mycobacterium tuberculosis identifies members of the ESAT-6 family as immunodominant T-cell antigens. Infect. Immun., 2000, vol. 68, no. 1, pp. 214–220. doi: 10.1128/IAI.68.1.214-220.2000
  15. WHO. BCG vaccine. WHO Wkly Epidemiol. Rec., 2004, vol. 79, no. 4, pp. 27–38.

Copyright (c) 2016 Dukhovlinov I.V., Fedorova E.A., Dobrovolskaya O.A., Bogomolova E.G., Chernyaeva E.N., Al-Shekhadat R.I., Simbirtsev A.S.

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.

This website uses cookies

You consent to our cookies if you continue to use our website.

About Cookies