Proteomic Analysis for Malonylastragaloside I in U937 Leukemia Cells by Modified Label-free Quantitative Strategy with LC Chip Q-TOF MS/MS

Abstract

Aim To study the differentially-expressed proteins for malonylastragaloside I in human leukemia U937 cells. Methods A suitable protein quantitative mode of label-free technique was established by LC Chip Q-TOF. It was found that the mode of peptide spectral intensity based on label-free quantification strategy of LC Chip Q-TOF showed high reproducibility, wide dynamic range, as well as simplicity and convenience. The optimized quantification mode of the peptide spectral intensity, normalization algorithm was applied to screen the differentially-expressed proteins for malonylastragaloside I (MA-I), a novel astragaloside from Astragalus sp., in human leukemia U937 cells. Results Fifteen differentially-expressed proteins with up or down-regulation over 2.0 folds were identified, six of which including HSC70, RPLP2, PHB2, PCNA, TCTP and ANXA11 were confirmed by Western blotting. We also observed significant activation of PARP1 and cleavage of caspases-3 and −9 by MA-I in U937 cells, and the pan-caspase inhibitor Z-VAD-FMK could reduce MA-I inhibited cell proliferation, indicating that caspase activation may be involved in MA-I induced cell toxicity. Conclusion This study has generated potentially functional proteins important for MA-I inhibited U937 cells proliferation by the optimized quantification mode of peptide spectral intensity.