title: Ligand-free palladium-mediated site-specific protein labeling inside gram-negative bacterial pathogens.
abstract: Palladium, a key transition metal in advancing modern organic synthesis, mediates diverse chemical conversions including many carbon-carbon bond formation reactions between organic compounds. However, expanding palladium chemistry for conjugation of biomolecules such as proteins, particularly within their native cellular context, is still in its infancy. Here we report the site-specific protein labeling inside pathogenic Gram-negative bacterial cells via a ligand-free palladium-mediated cross-coupling reaction. Two rationally designed pyrrolysine analogues bearing an aliphatic alkyne or an iodophenyl handle were first encoded in different enteric bacteria, which offered two facial handles for palladium-mediated Sonogashira coupling reaction on proteins within these pathogens. A GFP-based bioorthogonal reaction screening system was then developed, allowing evaluation of both the efficiency and the biocompatibilty of various palladium reagents in promoting protein-small molecule conjugation. The identified simple compound-Pd(NO3)2 exhibited high efficiency and biocompatibility for site-specific labeling of proteins in vitro and inside living E. coli cells. This Pd-mediated protein coupling method was further utilized to label and visualize a Type-III Secretion (T3S) toxin-OspF in Shigella cells. Our strategy may be generally applicable for imaging and tracking various virulence proteins within Gram-negative bacterial pathogens.
authors: Synthetic and Functional Biomolecules Center, Beijing National Laboratory for Molecular Sciences, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
authors: Jie
authors: J
authors: Li
authors: Shixian
authors: S
authors: Lin
authors: Jie
authors: J
authors: Wang
authors: Shang
authors: S
authors: Jia
authors: Maiyun
authors: M
authors: Yang
authors: Ziyang
authors: Z
authors: Hao
authors: Xiaoyu
authors: X
authors: Zhang
authors: Peng R
authors: PR
authors: Chen
mesh_terms: D001426:Bacterial Proteins / Q000032:analysis*; D004926:Escherichia coli / Q000166:cytology*; D049452:Green Fluorescent Proteins / Q000032:analysis; D010165:Palladium / Q000737:chemistry*; D012760:Shigella / Q000166:cytology*; D013194:Staining and Labeling / Q000379:methods
chemical_list: D001426:Bacterial Proteins; D049452:Green Fluorescent Proteins; D010165:Palladium