title: The on-bead digestion of protein corona on nanoparticles by trypsin immobilized on the magnetic nanoparticle.
abstract: Proteins interacting with nanoparticles would form the protein coronas on the surface of nanoparticles in biological systems, which would critically impact the biological identities of nanoparticles and/or result in the physiological and pathological consequences. The enzymatic digestion of protein corona was the primary step to achieve the identification of protein components of the protein corona for the bottom-up proteomic approaches. In this study, the investigation on the tryptic digestion of protein corona by the immobilized trypsin on a magnetic nanoparticle was carried out for the first time. As a comparison with the usual overnight long-time digestion and the severe self-digestion of free trypsin, the on-bead digestion of protein corona by the immobilized trypsin could be accomplished within 1h, along with the significantly reduced self-digestion of trypsin and the improved reproducibility on the identification of proteins by the mass spectrometry-based proteomic approach. It showed that the number of identified bovine serum (BS) proteins on the commercial Fe3O4 nanoparticles was increased by 13% for the immobilized trypsin with 1h digestion as compared to that of using free trypsin with even overnight digestion. In addition, the on-bead digestion of using the immobilized trypsin was further applied on the identification of human plasma protein corona on the commercial Fe3O4 nanoparticles, which leads the efficient digestion of the human plasma proteins and the identification of 149 human plasma proteins corresponding to putative critical pathways and biological processes.
authors.affiliation: CAS Key Laboratory of Separation Science for Analytical Chemistry, National Chromatographic R & A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences (CAS) , Dalian 116023, China; Graduate School of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Zhengyan
authors.initials: Z
authors.lastname: Hu
authors.affiliation: CAS Key Laboratory of Separation Science for Analytical Chemistry, National Chromatographic R & A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences (CAS) , Dalian 116023, China; Graduate School of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Zhao
authors.affiliation: CAS Key Laboratory of Separation Science for Analytical Chemistry, National Chromatographic R & A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences (CAS) , Dalian 116023, China; Graduate School of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Hongyan
authors.initials: H
authors.lastname: Zhang
authors.affiliation: CAS Key Laboratory of Separation Science for Analytical Chemistry, National Chromatographic R & A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences (CAS) , Dalian 116023, China; Graduate School of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: CAS Key Laboratory of Separation Science for Analytical Chemistry, National Chromatographic R & A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences (CAS) , Dalian 116023, China. Electronic address: wurenan@dicp.ac.cn.
authors.forename: Ren'an
authors.initials: R
authors.lastname: Wu
authors.affiliation: CAS Key Laboratory of Separation Science for Analytical Chemistry, National Chromatographic R & A Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences (CAS) , Dalian 116023, China. Electronic address: hanfazou@dicp.ac.cn.
authors.forename: Hanfa
authors.initials: H
authors.lastname: Zou
mesh_terms: D000818:Animals; D001798:Blood Proteins / Q000032:analysis / Q000378:metabolism; D002417:Cattle; D004800:Enzymes, Immobilized / Q000378:metabolism*; D006801:Humans; D058185:Magnetite Nanoparticles / Q000648:ultrastructure; D040901:Proteomics; D015203:Reproducibility of Results; D012710:Serum Albumin, Bovine / Q000378:metabolism; D053719:Tandem Mass Spectrometry; D014357:Trypsin / Q000378:metabolism*
chemical_list: D001798:Blood Proteins; D004800:Enzymes, Immobilized; D058185:Magnetite Nanoparticles; D012710:Serum Albumin, Bovine; D014357:Trypsin
keywords: Digestion; Fe(3)O(4); Immobilized trypsin; Magnetic nanoparticle; Plasma; Protein corona