Integration of fluorescence imaging with proteomics enables visualization and identification of metallo-proteomes in living cells.
Metalloproteins account for nearly one-third of proteins in proteomes. To date, the identification of metalloproteins relies mainly on protein purification and the subsequent characterization of bound metals, which often leads to losses of metal ions bound weakly and transiently. Herein, we developed a strategy to visualize and subsequently identify endogenous metalloproteins and metal-binding proteins in living cells via integration of fluorescence imaging with proteomics. We synthesized a "metal-tunable" fluorescent probe (denoted as Mn+-TRACER) that rapidly enters cells to target proteins with 4-40 fold fluorescence enhancements. By using Ni2+-TRACER as an example, we demonstrate the feasibility of tracking Ni2+-binding proteins in vitro, while cellular small molecules exhibit negligible interference on the labeling. We identified 44 Ni2+-binding proteins from microbes using Helicobacter pylori as a showcase. We further applied Cu2+-TRACER to mammalian cells and found 54 Cu2+-binding proteins. The strategy we report here provides a great opportunity to track various endogenous metallo-proteomes and to mine potential targets of metallodrugs.
Department of Chemistry, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, P. R. China. hsun@hku.hk.
Yau-Tsz
YT
Lai
Department of Chemistry, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, P. R. China. hsun@hku.hk.
Ya
Y
Yang
Department of Chemistry, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, P. R. China. hsun@hku.hk.
Ligang
L
Hu
Department of Chemistry, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, P. R. China. hsun@hku.hk.
Tianfan
T
Cheng
Department of Chemistry, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, P. R. China. hsun@hku.hk.
Yuen-Yan
YY
Chang
Department of Chemistry, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, P. R. China. hsun@hku.hk.
Mohamad
M
Koohi-Moghadam
MOE Key Laboratory of Bioinorganic and Synthetic Chemistry, School of Chemistry and Chemical Engineering, Sun Yat-sen University, Guangzhou, P. R. China.
Yuchuan
Y
Wang
Department of Chemistry, The Chinese University of Hong Kong, Hong Kong, P. R. China.
Jiang
J
Xia
Center for Individualized Medicine & Department of Health Sciences Research, Mayo Clinic, Scottsdale, AZ 85259 USA and Department of Biomedical Informatics, Arizona State University, Scottsdale, AZ 85259 USA.
Junwen
J
Wang
Department of Chemistry, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, P. R. China. hsun@hku.hk.
Hongyan
H
Li
Department of Chemistry, The University of Hong Kong, Pokfulam Road, Hong Kong SAR, P. R. China. hsun@hku.hk.
Hongzhe
0000-0001-6697-6899
H
Sun
D002352:Carrier Proteins / Q000378:metabolism*; D005453:Fluorescence; D006367:HeLa Cells; D006801:Humans; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D008667:Metalloproteins / Q000378:metabolism*; D008670:Metals / Q000378:metabolism*; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D013050:Spectrometry, Fluorescence
D002352:Carrier Proteins; D008667:Metalloproteins; D008670:Metals; D020543:Proteome