title: Vortex-assisted tryptic digestion.
abstract: The effect of vortex-induced vibration during tryptic digestion was investigated by applying different vibrational speeds (0, 600, 1200, or 2500 rpm) to digestion solutions for varying durations (10, 20, 30, 40, or 60 min) at two different incubation temperatures (25°C or 37°C). The most rapid digestion was observed with the highest vibrational speed and temperature. With the application of 2500 rpm at 37°C, the tryptic digestion of each of three standard proteins (cytochrome c, myoglobin, or bovine serum albumin) provided complete disappearance of the protein within 60 min, as determined by matrix-assisted laser desorption/ionization mass spectrometry. Compared to conventional overnight digestion, 60-min vortex-assisted tryptic digestion generated longer peptides, due primarily to the limited digestion time and provided better sequence coverages (89% vs. 78% for cytochrome c, 100% vs. 87% for myoglobin, and 38% vs. 26% for BSA). The longer peptides should be advantageous to analytical methods such as the middle-down approach that benefit from increased sequence coverage of proteins. Vortex-assisted tryptic digestion is expected to be a useful method for rapid tryptic digestion of proteins.
authors.affiliation: Department of Chemistry, Chungnam National University, Daejeon, South Korea.
authors.forename: Hyo-Jik
authors.initials: HJ
authors.lastname: Yang
authors.forename: Seongjae
authors.initials: S
authors.lastname: Shin
authors.forename: Jinhee
authors.initials: J
authors.lastname: Kim
authors.forename: Jangmi
authors.initials: J
authors.lastname: Hong
authors.forename: Sunyoung
authors.initials: S
authors.lastname: Lee
authors.forename: Jeongkwon
authors.initials: J
authors.lastname: Kim
mesh_terms: D000818:Animals; D002417:Cattle; D045304:Cytochromes c / Q000737:chemistry / Q000378:metabolism; D009211:Myoglobin / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013997:Time Factors; D014357:Trypsin / Q000737:chemistry* / Q000378:metabolism*; D014732:Vibration
chemical_list: D009211:Myoglobin; D010446:Peptide Fragments; D012710:Serum Albumin, Bovine; D045304:Cytochromes c; D014357:Trypsin