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Abstract:

A slab gel electrophoresis apparatus with the ability to operate over a pressure range of 10-3 to 2 kbar is described. The system presented here is an improvement of a previous apparatus (A. A. Paladini, J. L. Silva, and G. Weber, Anal. Biochem. 161, 358-364, 1987). It consists of a flat bed gel, with a significantly enlarged buffer reservoir, which eliminates the requirement of high concentrations of running buffers, and at the same time allows shorter runs, leading to enhanced resolution and reproducibility. The application of the method to the dissociation of the tetramer glycogen phosphorylase a as a function of hydrostatic pressure is described. The flat geometry of the apparatus allows for the first time the analysis of the stability of oligomers and their constituent subunits to chemical denaturation by urea gradient electrophoresis gels at high pressure. Dimeric hexokinase shows a reversible cooperative unfolding transition with a midpoint at 3.8 M urea. In contrast, the monomers unfold at very low urea concentration (<1.0 M). The observed differences in stability validates oligomerization as an important stabilizing element of the protein structure. © 1994 Academic Press, Inc.

Registro:

Documento: Artículo
Título:Analysis of dissociation and unfolding of oligomeric proteins using a flat bed gel electrophoresis at high pressure
Autor:Paladini, A.A.; Weber, G.; Erijmant, L.
Filiación:Institute de Investigaciones en Ingenieria Genetica y Biologia Molecular, Facultad de Ciencias Exactas y Naturales (UBA), 1428 Buenos Aires, Argentina
Department of Biochemistry, University of Illinois at Urbana-Champaign, Urbana, IL 61801, United States
Año:1994
Volumen:218
Número:2
Página de inicio:364
Página de fin:369
DOI: http://dx.doi.org/10.1006/abio.1994.1193
Título revista:Analytical Biochemistry
Título revista abreviado:ANAL. BIOCHEM.
ISSN:00032697
Registro:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00032697_v218_n2_p364_Paladini

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Citas:

---------- APA ----------
Paladini, A.A., Weber, G. & Erijmant, L. (1994) . Analysis of dissociation and unfolding of oligomeric proteins using a flat bed gel electrophoresis at high pressure. Analytical Biochemistry, 218(2), 364-369.
http://dx.doi.org/10.1006/abio.1994.1193
---------- CHICAGO ----------
Paladini, A.A., Weber, G., Erijmant, L. "Analysis of dissociation and unfolding of oligomeric proteins using a flat bed gel electrophoresis at high pressure" . Analytical Biochemistry 218, no. 2 (1994) : 364-369.
http://dx.doi.org/10.1006/abio.1994.1193
---------- MLA ----------
Paladini, A.A., Weber, G., Erijmant, L. "Analysis of dissociation and unfolding of oligomeric proteins using a flat bed gel electrophoresis at high pressure" . Analytical Biochemistry, vol. 218, no. 2, 1994, pp. 364-369.
http://dx.doi.org/10.1006/abio.1994.1193
---------- VANCOUVER ----------
Paladini, A.A., Weber, G., Erijmant, L. Analysis of dissociation and unfolding of oligomeric proteins using a flat bed gel electrophoresis at high pressure. ANAL. BIOCHEM. 1994;218(2):364-369.
http://dx.doi.org/10.1006/abio.1994.1193