INVESTIGADORES
CESARI Andreina
artículos
Título:
A simple technique to improve the resolution of membrane acidic proteins of the haloarchaeon Haloferax volcanii by 2D electrophoresis
Autor/es:
PAGGI, R; GIMENEZ, MI; DE CASTRO, R; CESARI, A
Revista:
Electrophoresis
Editorial:
WILEY-VCH Verlag GmbH & Co
Referencias:
Lugar: Weinheim; Año: 2014 vol. 35 p. 3518 - 3522
ISSN:
1522-2683
Resumen:
Proteins present in the archaeal cell envelope play key roles in a variety of processes necessary for survival in extreme environments. The haloarchaeon Haloferax volcanii is a good model for membrane proteomic studies because its genome sequence is known, it can be genetically manipulated and a number of studies at the ?omics? level have been performed in this organism. This work reports an easy strategy to improve the resolution of acidic membrane proteins from H. volcanii by 2DE. The method is based on the solubilization, delipidation and salt removal from membrane proteins. Due to the abundance of the S-layer glycoprotein in membrane protein extracts, other proteins from the envelope are consequently underrepresented. Thus, a protocol to reduce the amount of the S-layer glycoprotein by EDTA treatment was applied and 11 cm narrow range pH (3.9 ? 5.1) IPG strips were used to separate the remaining proteins. Using this method, horizontal streaking was substantially decreased and at least 75 defined spots (20% of the predicted membrane proteome within this pI/Mw range) were reproducibly detected. Removal of the S-layer glycoprotein from membrane protein extracts can be applied to increase protein load for 2DE as well as for other proteomic methods.