Limitations to the comparative proteomic analysis of thrombopoietin producing Chinese hamster ovary cells treated with sodium butyrate.
Sodium butyrate (NaBu) is known to enhance the specific productivity of Chinese hamster ovary cells expressing human thrombopoietin. In order to better understand the intracellular responses of these cells resulting from NaBu treatment, the proteomic profiles of cells treated with various concentrations of NaBu (0-3mM) were compared using two-dimensional electrophoresis (2-DE). Based on spot intensities, 80 high intensity protein spots were selected. Fifty-six of the 80 protein spots, which represent 28 different kinds of proteins, were identified by MALDI-TOF-MS and MS/MS. Compared to control without NaBu treatment, the expression levels of 2 proteins (glucose regulated protein 78 (GRP 78) and peroxiredoxin 4) were increased over two fold with NaBu treatment and the expression level of phosphopyruvate hydratase was decreased over two fold with NaBu treatment. Due to multiplicity (multiple spots for one protein), a change in one single spot intensity from a 2-DE gel image may not represent the total change in expression level for that protein. Western blot analyses of GRP78, HSC70 and ERp57 confirmed the results of the MS analyses. However, a degree of change in expression level differed between the two methods, suggesting the necessity of a validating method to determine the total amount of the protein.
Baik
Jong Youn
JY
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 373-1 Kusong-Dong, Yusong-Gu, Daejon 305-701, Republic of Korea.
Joo
Eun Jin
EJ
Kim
Young Hwan
YH
Lee
Gyun Min
GM
D000818:Animals; D015153:Blotting, Western; D002087:Butyrates / Q000494:pharmacology*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D015180:Electrophoresis, Gel, Two-Dimensional; D000091342:Endoplasmic Reticulum Chaperone BiP; D005786:Gene Expression Regulation / Q000187:drug effects; D020543:Proteome / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D053719:Tandem Mass Spectrometry; D013926:Thrombopoietin / Q000032:analysis / Q000378:metabolism*
D002087:Butyrates; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D020543:Proteome; D013926:Thrombopoietin