Producción y purificación de osteopontina bovina recombinante mediante Escherichia coli como fábrica celular

Vista sencilla de ítem
dc.audienceInvestigador
dc.coverageColombia
dc.creatorBrijaldo Villamizar, Angela Patricia
dc.creatorLondoño Méndez, María Camila
dc.creatorArbeláez Ramírez, Luis Fernando
dc.creatorRueda Alfonso, Fabian Leonardo
dc.date2024-01-18T15:51:42Z
dc.date2024-01-18T15:51:42Z
dc.date2022-05-03
dc.date2022
dc.date.accessioned2026-06-27T04:37:16Z
dc.descriptionLos programas de reproducción y mejoramiento animal requieren la optimización de herramientas biotecnológicas capaces de favorecer los índices reproductivos en diversas especies. El uso de aditivos proteicos que mejoren la criopreservación espermática y la producción de embriones in vitro, parece ser una alternativa interesante. La Osteopontina se ha relacionado con el potencial fecundante del espermatozoide y con el desarrollo embrionario temprano. El objetivo de este trabajo fue determinar las condiciones óptimas para la producción de Osteopontina recombinante (rOPN) mediante el uso de Escherichia coli como fábrica celular. Para esto, el gen de la OPN se insertó en un vector de expresión pET28(a+) inducible por IPTG, con resistencia a la Kanamicina y una cola de histidinas (6xHis-tag). El constructo resultante se usó para transformar células competentes de E. Coli BL21-StarTM. Las colonias transformadas se usaron para la producción de rOPN-H6 a 20, 30 y 37 °C, probándose dos concentraciones del inductor IPTG (1.0 y 0.1mM). Se realizó una purificación de la rOPN-H6 mediante columnas de afinidad con imidazol (10, 50, 200, 350, 500mM). Los resultados evidenciaron que la producción de rOPN-H6 solo fue exitosa a 37°C independiente de la concentración de IPTG empleada. La purificación de la rOPN-H6 fue exitosa usando imidazol a 200mM, con una aparente tendencia a la dimerización luego de obtener la proteína purificada. De este modo, se concluye cuáles son las mejores condiciones para obtener la OPN recombinante, sugiriendo su potencial uso en ensayos de criopreservación espermática y en medios de cultivo para producción de embriones in vitro.
dc.descriptionGanadería bovina
dc.formatapplication/pdf
dc.formatapplication/pdf
dc.identifierhttps://revistas.uptc.edu.co/index.php/ciencia_agricultura/article/view/14071
dc.identifier0122-8420
dc.identifierhttp://hdl.handle.net/20.500.12324/38763
dc.identifier10.19053/01228420.v19.n2.2022.14071
dc.identifierreponame:Biblioteca Digital Agropecuaria de Colombia
dc.identifierinstname:Corporación colombiana de investigación agropecuaria AGROSAVIA
dc.identifier.urihttp://hdl.handle.net/123456789/33301
dc.languagespa
dc.publisherUniversidad Pedagógica y Tecnológica de Colombia
dc.relationRevista Ciencia y Agricultura
dc.relation19
dc.relation2
dc.relation47
dc.relation63
dc.relationAlvarez-gallardo, H., Kjelland, M. E., Moreno, J. F., Jr, T. H. W., Lara-sagaho, A. V, Randel, R. D., Lammoglia, M. A., Pe, M., Romo, S., & Espero, E. (2013). Gamete Therapeutics : Recombinant Protein Adsorption by Sperm for Increasing Fertility via Artificial Insemination. 8(6). https://doi.org/10.1371/journal.pone.0065083.
dc.relationBarrios, B., Pe, R., Gallego, M., Muin, T., & Cebria, A. (2000). Seminal Plasma Proteins Revert the Cold-Shock Damage on Ram Sperm Membrane 1 and Jose. 1537, 1531–1537. https://doi.org/10.1095/biolreprod63.5.1531
dc.relationBoccia, L., Di Francesco, S., Neglia, G., De Blasi, M., Longobardi, V., Campanile, G., & Gasparrini, B. (2013). Osteopontin improves sperm capacitation and in vitro fertilization efficiency in buffalo (Bubalus bubalis). Theriogenology, 80(3), 212– 217. https://doi.org/10.1016/j.theriogenology.2013.04.017.
dc.relationBradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72(1–2), 248–254. https://doi.org/10.1016/0003-2697(76)90527-3.
dc.relationBustamanete Filho, I. C. (2010). Cloning, expression and purification of proteins from bovine seminal plasma related with semen freezability. In Science. e [Trabajo de grado, Repositorio institucional Universidade Federal do Rio Grande do sul. https://lume.ufrgs.br/handle/10183/23020
dc.relationBustamante-Filho, I. C., Renato Menegassi, S., Ribas Pereira, G., Dias Salton, G., Mosena Munari, F., Roberto Schneider, M., Costa Mattos, R., Otávio Jardim Barcellos, J., Pereira Laurino, J., Obino Cirne-Lima, E., & Inês Mascarenhas Jobim, M. (2021). Bovine seminal plasma osteopontin: Structural modelling, recombinant expression and its relationship with semen quality. Andrologia, 53(1), 1–15. https://doi.org/10.1111/and.13905
dc.relationBustamante-Filho, I., Salton, G. D., Munari, F. M., Schneider, M. R., Mattos, R. C., Laurino, J. P., Cirne-Lima, E. O., & Jobim, M. I. M. (2014). Recombinant expression and purification of the bovine acidic Seminal Fluid Protein. Animal Reproduction, 11(2), 96–103.
dc.relationCajazeiras, J. B., Melo, L. M., Albuquerque, E. S., Rádis-Baptista, G., Cavada, B. S., & Freitas, V. J. F. (2009). Analysis of protein expression and a new prokaryotic expression system for goat (Capra hircus) spermadhesin Bdh-2 cDNA. Genetics and Molecular Research : GMR, 8(3), 1147–1157. https://doi.org/10.4238/ vol8-3gmr639
dc.relationCancel, a M., Chapman, D. a, & Killian, G. J. (1997). Osteopontin is the 55-kilodalton fertility-associated protein in Holstein bull seminal plasma. Biology of Reproduction, 57(6), 1293–1301. https://doi.org/10.1095/biolreprod57.6.1293
dc.relationCarbonell, X., & Villaverde, A. (2002). Protein aggregated into bacterial inclusion bodies does not result in protection from proteolytic digestion. 1939–1944. https://doi.org/10.1023/A:1021173114193
dc.relationEduardo, C., Souza, A., Moura, A. A., & Killian, G. J. (2008). Binding patterns of bovine seminal plasma proteins A1 / A2 , 30 kDa and osteopontin on ejaculated sperm before and after incubation with isthmic and ampullary oviductal fluid ଝ, ଝଝ. 105, 72–89. https://doi.org/10.1016/j.anireprosci.2007.11.027
dc.relationGabler, C., Chapman, D. A., & Killian, G. J. (2003). Expression and presence of osteopontin and integrins in the bovine oviduct during the oestrous cycle. Reproduction, 126(6), 721–729. https://doi.org/10.1530/rep.0.1260721
dc.relationGarcía-Fraga, B., da Silva, A. F., López-Seijas, J., & Sieiro, C. (2015). Optimized expression conditions for enhancing production of two recombinant chitinolytic enzymes from different prokaryote domains. Bioprocess and Biosystems Engineering, 38(12), 2477–2486. https://doi.org/10.1007/s00449-015-1485-5
dc.relationHaglin, E. R., Yang, W., Briegel, A., & Thompson, L. K. (2017). His-Tag-Mediated Dimerization of Chemoreceptors Leads to Assembly of Functional Nanoarrays. Biochemistry, 56(44), 5874–5885. https://doi.org/10.1021/acs. biochem.7b00698
dc.relationIcer, M. A., & Gezmen-Karadag, M. (2018). The multiple functions and mechanisms of osteopontin. Clinical Biochemistry, 59(July), 17–24. https://doi.org/10.1016/j. clinbiochem.2018.07.003
dc.relationLiu, Q., Xie, Q. Z., Zhou, Y., & Yang, J. (2014). Osteopontin is expressed in the oviduct and promotes fertilization and preimplantation embryo development of mouse. Zygote, 760. https://doi.org/10.1017/S0967199414000483
dc.relationMonaco, E., Gasparrini, B., Boccia, L., De Rosa, A., Attanasio, L., Zicarelli, L., & Killian, G. (2009). Effect of osteopontin (OPN) on in vitro embryo development in cattle. Theriogenology, 71(3), 450–457. https://doi.org/10.1016/j. theriogenology.2008.08.012
dc.relationMoura, A. A. (2005). Seminal plasma proteins and fertility indexes in the bull : The case for osteopontin. Anim Reprod, 2, March, 3–10.
dc.relationNature. (1970). Laemmli buffer Background Purpose of the Laemmli buffer Laemmli Buffer Recipe Preparation Recommended Storage Temperature of Laemmli buffer Laemmli buffer References. Nature.
dc.relationOno, B., Kimiduka, H., Kubota, M., Okuno, K., & Yabuta, M. (2007). Role of the ompT mutation in stimulated decrease in colony-forming ability due to intracellular protein aggregate formation in Escherichia coli strain BL21. Bioscience, Biotechnology, and Biochemistry, 71(2), 504–512. https://doi.org/10.1271/bbb.60541
dc.relationPeris, S. I., Bilodeau, J. F., Dufour, M., & Bailey, J. L. (2007). Impact of cryopreservation and reactive oxygen species on DNA integrity, lipid peroxidation, and functional parameters in ram sperm. Molecular Reproduction and Development, 74(7), 878–892. https://doi.org/10.1002/mrd.20686
dc.relationPrince, C. W. (1989). Secondary structure predictions for rat osteopontin. Connective Tissue Research, 21(1–4), 15–20. https://doi.org/10.3109/03008208909049991
dc.relationRetnoningrum, D. S., Pramesti, H. T., Santika, P. Y., Valerius, O., Asjarie, S., & Suciati, T. (2012). Codon optimization for high level expression of human bone morphogenetic protein-2 in Escherichia coli. Protein Expression and Purification, 84(2), 188–194. https://doi.org/10.1016/j.pep.2012.05.010
dc.relationRueda, F., Cano-Garrido, O., Mamat, U., Wilke, K., Seras-Franzoso, J., García-Fruitós, E., & Villaverde, A. (2014). Production of functional inclusion bodies in endotoxin-free Escherichia coli. Applied Microbiology and Biotechnology, 98(22), 9229– 9238. https://doi.org/10.1007/s00253-014-6008-9
dc.relationRueda, F., Céspedes, M. V., Conchillo-Solé, O., Sánchez-Chardi, A., Seras-Franzoso, J., Cubarsi, R., Gallardo, A., Pesarrodona, M., Ferrer-Miralles, N., Daura, X., Vázquez, E., García-Fruitós, E., Mangues, R., Unzueta, U., & Villaverde, A. (2015). Bottom-Up Instructive Quality Control in the Biofabrication of Smart Protein Materials. Advanced Materials, 27(47), 7816–7822. https://doi.org/10.1002/ adma.201503676
dc.relationRueda, F., Garcés P, T., Herrera L, R., Arbeláez R, L., Peña J, M., Velásquez P, H., Hernández V, A., & Cardozo C, J. (2013). Seminal plasma proteins increase the post-thaw sperm viability of Sanmartinero bull’s semen. Revista MVZ Cordoba, 18(1), 3327–3335.
dc.relationSicherle, C. C., Maia, M. S., Bicudo, S. D., Rodello, L., & Azevedo, H. C. (2011). Lipid peroxidation and generation of hydrogen peroxide in frozen-thawed ram semen supplemented with catalase or Trolox. Small Ruminant Research, 95(2–3), 144– 149. https://doi.org/10.1016/j.smallrumres.2010.10.011
dc.relationSingh, B. P., Asthana, A., Basu, A., & Tangirala, R. (2020). Conserved core tryptophans of FnII domains are crucial for the membranolytic and chaperone-like activities of bovine seminal plasma protein PDC-109. 594, 509–518. https://doi. org/10.1002/1873-3468.13617
dc.relationSivashanmugam, A., Murray, V., Cui, C., Zhang, Y., Wang, J., & Li, Q. (2009). Practical protocols for production of very high yields of recombinant proteins using Escherichia coli. 18(1), 936–948. https://doi.org/10.1002/pro.102
dc.relationThérien, I., Moreau, R., & Manjunath, P. (1999). Bovine seminal plasma phospholipid-binding proteins stimulate phospholipid efflux from epididymal sperm. Biology of Reproduction, 61, 590–598. https://doi.org/10.1095/biolreprod61.3.590
dc.relationWatson, P. F. (1995). Recent developments and concepts in the cryopreservation of spermatozoa and the assessment of their post-thawing function. Reproduction, Fertility and Development, 7(4), 871–891. https://doi.org/10.1071/RD9950871
dc.relationWei, R., Wong, J. P. C., & Kwok, H. F. (2017). Osteopontin - A promising biomarker for cancer therapy. Journal of Cancer, 8(12), 2173–2183. https://doi.org/10.7150/ jca.20480
dc.relationWillforss, J., Morrell, J. M., Resjö, S., Hallap, T., Padrik, P., Siino, V., de Koning, D. J., Andreasson, E., Levander, F., & Humblot, P. (2021). Stable bull fertility protein markers in seminal plasma. Journal of Proteomics, 236, 104135. https://doi. org/10.1016/j.jprot.2021.104135
dc.rightsAttribution-ShareAlike 4.0 International
dc.rightshttp://creativecommons.org/licenses/by-sa/4.0/
dc.sourceCiencia y Agricultura; Vol. 19, Núm. 2 (2022):Ciencia y Agricultura (Mayo);p. 47 -63.
dc.subjectGenética y mejoramiento animal - L10
dc.subjectReproducción animal
dc.subjectProteínas recombinantes
dc.subjectSemen
dc.subjectCriopreservación
dc.subjectGanadería y especies menores
dc.subjecthttp://aims.fao.org/aos/agrovoc/c_5d5904a8
dc.subjecthttp://aims.fao.org/aos/agrovoc/c_36920
dc.subjecthttp://aims.fao.org/aos/agrovoc/c_6961
dc.subjecthttp://aims.fao.org/aos/agrovoc/c_24155
dc.thumbnailhttps://repository.agrosavia.co/bitstreams/b14aab3e-0061-491a-b69c-02e270126be4/download
dc.titleProducción y purificación de osteopontina bovina recombinante mediante Escherichia coli como fábrica celular
dc.titleProduction and purification of recombinant bovine osteopontin using Escherichia coli as a Cell Factory
dc.typeArtículo científico

Archivos

Colecciones

Científico