Nitrilase-Activatable Noncanonical Amino Acid Precursors for Cell-Selective Metabolic Labeling of Proteomes.
Cell-selective protein metabolic labeling is of great interest for studying cell-cell communications and tissue homeostasis. We herein describe a nitrilase-activatable noncanonical amino acid tagging (NANCAT) strategy that exploits an exogenous nitrilase to enzymatically convert the nitrile-substituted precursors to their corresponding noncanonical amino acids (ncAAs), l-azidohomoalanine (Aha) or homopropargylglycine (Hpg), in living cells. Only cells expressing the nitrilase can generate Aha or Hpg in cellulo and metabolically incorporate them into the nascent proteins. Subsequent click-labeling of the azide- or alkyne-incorporated proteins with fluorescent probes or with affinity tags enables visualization and proteomic profiling of nascent proteomes, respectively. We have demonstrated that NANCAT can serve as a versatile strategy for cell-selective labeling of proteomes in both bacterial and mammalian cells.
College of Chemistry and Molecular Engineering, ‡Academy for Advanced Interdisciplinary Studies, §Peking-Tsinghua Center for Life Sciences, ∥Synthetic and Functional Biomolecule Center, and ⊥Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Peking University , Beijing, 100871, China.
Zefan
Z
Li
College of Chemistry and Molecular Engineering, ‡Academy for Advanced Interdisciplinary Studies, §Peking-Tsinghua Center for Life Sciences, ∥Synthetic and Functional Biomolecule Center, and ⊥Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Peking University , Beijing, 100871, China.
Yuntao
Y
Zhu
College of Chemistry and Molecular Engineering, ‡Academy for Advanced Interdisciplinary Studies, §Peking-Tsinghua Center for Life Sciences, ∥Synthetic and Functional Biomolecule Center, and ⊥Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Peking University , Beijing, 100871, China.
Yuting
Y
Sun
College of Chemistry and Molecular Engineering, ‡Academy for Advanced Interdisciplinary Studies, §Peking-Tsinghua Center for Life Sciences, ∥Synthetic and Functional Biomolecule Center, and ⊥Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Peking University , Beijing, 100871, China.
Ke
K
Qin
College of Chemistry and Molecular Engineering, ‡Academy for Advanced Interdisciplinary Studies, §Peking-Tsinghua Center for Life Sciences, ∥Synthetic and Functional Biomolecule Center, and ⊥Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Peking University , Beijing, 100871, China.
Weibing
W
Liu
College of Chemistry and Molecular Engineering, ‡Academy for Advanced Interdisciplinary Studies, §Peking-Tsinghua Center for Life Sciences, ∥Synthetic and Functional Biomolecule Center, and ⊥Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Peking University , Beijing, 100871, China.
Wen
W
Zhou
College of Chemistry and Molecular Engineering, ‡Academy for Advanced Interdisciplinary Studies, §Peking-Tsinghua Center for Life Sciences, ∥Synthetic and Functional Biomolecule Center, and ⊥Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Peking University , Beijing, 100871, China.
Xing
X
Chen
D000409:Alanine / Q000031:analogs & derivatives* / Q000378:metabolism; D000480:Alkynes / Q000378:metabolism*; D000619:Aminohydrolases / Q000378:metabolism*; D000818:Animals; D016466:CHO Cells; D003412:Cricetulus; D005998:Glycine / Q000031:analogs & derivatives* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D020543:Proteome / Q000032:analysis* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D013194:Staining and Labeling
D000480:Alkynes; D020543:Proteome; C506098:azidohomoalanine; C551041:homopropargylglycine; D000619:Aminohydrolases; C021867:nitrilase; D000409:Alanine; D005998:Glycine