Chinese Journal of Tissue Engineering Research ›› 2010, Vol. 14 ›› Issue (50): 9374-.doi: 10.3969/j.issn.1673-8225.2010.50.016

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Methods for proteasome activity measurement and component analysis  

Chen Xiao-qin1, Pan Xiao-fen2, Xie Jun2   

  1. 1 Department of Hematology, Cancer Hospital, Sun Yat-sen University, Guangzhou  510080, Guangdong Province, China; 2 Guangzhou Institute of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou  510663, Guangdong Province, China
  • Online:2010-12-10 Published:2010-12-10
  • About author:Chen Xiao-qin★, Master, Department of Hematology, Cancer Hospital, Sun Yat-sen University, Guangzhou 510080, Guangdong Province, China xiaoqinchen2005@ hotmail.com
  • Supported by:

    the Natural Science Foundation of Guangdong Province, No. 06107503*

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Abstract:

BACKGROUND: 26S proteasome plays an important role in maintaining normal cell cycle, proliferation, and survival. However, there is not a precise method can test in vivo proteasome activity and composition. 
OBJECTIVE: To establish methods for analyzing proteasome activity and composition in cancer tissues.
METHODS: Three specific fluorogenic peptide substrates: Suc-LLVY-AMC, Z-ARR-AMC and Z-LLE-AMC were used to monitor chymotrypsin-like, trypsin-like and peptidly-glutamyl peptide-hydrolyzing-like (PGPH-like) proteolytic activity, and a proteasome-specific affinity probe AdaK(Bio)Ahx3L3VS was employed to assay the β1/β1i, β2/β2i, and β5/β5i subunits of proteasome.
RESULTS AND CONCLUSION: We investigated the catalytic activity and the expression of the component of proteasome, and the effects of proteasome inhibitor PS341 and PSI on the activity and composition of proteasome in erythroleukemia K562 cells. Our results showed that the expression of the β2 and β5 subunits of proteasome was high, while PS341 and PSI inhibited β5/β5i subunits, and the chymotrypsin-like activity of proteasome in K562 cells. Methods for analyzing proteasome activity and composition was established and supplied experimental methods for identifying in vitro activity of proteasome inhibitors.

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