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Application of free‐flow IEF to identify protein candidates changing under microgravity conditions
Author(s) -
Pietsch Jessica,
Kussian Richard,
Sickmann Albert,
Bauer Johann,
Weber Gerhard,
Nissum Mikkel,
Westphal Kriss,
Egli Marcel,
Grosse Jirka,
Schönberger Johann,
Wildgruber Robert,
Infanger Manfred,
Grimm Daniela
Publication year - 2010
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200900226
Subject(s) - proteome , two dimensional gel electrophoresis , gel electrophoresis , chemistry , proteomics , microbiology and biotechnology , free flow electrophoresis , cell culture , chromatography , polyacrylamide gel electrophoresis , biochemistry , biology , gel electrophoresis of proteins , genetics , gene , enzyme
Using antibody‐related methods, we recently found that human thyroid cells express various proteins differently depending on whether they are cultured under normal gravity (1g) or simulated microgravity (s‐μg). In this study, we performed proteome analysis in order to identify more gravity‐sensitive thyroid proteins. Cells cultured under 1g or s‐μg conditions were sonicated. Proteins released into the supernatant and those remaining in the cell fragments were fractionated by free‐flow IEF. The fractions obtained were further separated by SDS‐gel electrophoresis. Selected gel pieces were excised and their proteins were determined by MS. A total of 235 different proteins were found. Out of 235 proteins, 37 appeared to be first identifications in human thyroid cells. Comparing SDS gel lanes of equally numbered free‐flow IEF fractions revealed similar patterns with a number of identical bands if proteins of a distinct cell line had been applied, irrespective of whether the cells had been cultured under 1g or s‐μg. Most of the identical band pairs contained identical proteins. However, the concentrations of some types of proteins were different within the two pieces of gel. Proteins that concentrated differently in such pieces of gel are considered as candidates for further investigations of gravitational sensitivity.