title: A nanoporous reactor for efficient proteolysis.
abstract: A nanoreactor based on mesoporous silicates is described for efficient tryptic digestion of proteins within the mesochannels. Cyano-functionalized mesoporous silicate (CNS), with an average pore diameter of 18 nm, is a good support for trypsin, with rapid in situ digestion of the model proteins, cytochrome c and myoglobin. The generated peptides were analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Proteolysis by trypsin-CNS is much more efficient than in-solution digestion, which can be attributed to nanoscopic confinement and concentration enrichment of the substrate within the mesopores. Proteins at concentrations of 2 ng muL(-1) were successfully identified after digestion for 20 min. A biological complex sample extracted from the cytoplasm of human liver tissue was digested by using the CNS-based reactor. Coupled with reverse-phase HPLC and MALDI-TOF MS/MS, 165 proteins were identified after standard protein data searching. This nanoreactor combines the advantages of short digestion time with retention of enzymatic activity, providing a promising way to advance the development of proteomics.
authors.affiliation: Department of Chemistry, Institute of Biomedical Sciences, Fudan University, Shanghai 200433, China.
authors.forename: Liang
authors.initials: L
authors.lastname: Qiao
authors.forename: Yun
authors.initials: Y
authors.lastname: Liu
authors.forename: Sarah P
authors.initials: SP
authors.lastname: Hudson
authors.forename: Pengyuan
authors.initials: P
authors.lastname: Yang
authors.forename: Edmond
authors.initials: E
authors.lastname: Magner
authors.forename: Baohong
authors.initials: B
authors.lastname: Liu
mesh_terms: D045304:Cytochromes c / Q000378:metabolism*; D006454:Hemoglobins / Q000378:metabolism; D006801:Humans; D013058:Mass Spectrometry; D008970:Molecular Weight; D049329:Nanostructures / Q000737:chemistry*; D016062:Porosity; D014357:Trypsin / Q000378:metabolism*
chemical_list: D006454:Hemoglobins; D045304:Cytochromes c; D014357:Trypsin