Mass spectrometry of the human pituitary proteome: identification of selected proteins

Abstract

The field of proteomics involves the combined application of advanced separation techniques, mass spectrometry, and bioinformatics tools to characterize proteins in complex biological mixtures. Here we report the identification of nine proteins from the human pituitary proteome, using the proteomics approach. The pituitary proteins were separated by two-dimensional electrophoresis, and were visualized by silver staining. The proteins of interest were subjected to in-gel digestion with trypsin, and the masses of the resulting peptides were determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. This tryptic mass map was used to identify the proteins through a search of a protein-sequence database. The identified proteins include important hormones, and enzymes with various catalytic activities. These proteins will be used to construct a two-dimensional reference database of the human pituitary. This database will be employed to study changes in the pituitary proteome that are associated with the formation of pituitary tumors.

Sulfolobus tengchongensis Spindle-Shaped Virus STSV1: Virus-Host Interactions and Genomic Features

“…For protein identification, gel slices containing protein bands were excised from the stained gel. In-gel digestion of the proteins was performed as described previously (10). The digests were dissolved in 0.1% formic acid, and purified with ZipTip C18 pipette tips (Millipore, Bedford, MA) according to the manufacturer's procedure.…”

Antibodies with Dual Reactivity to Plasminogen and Complementary PR3 in PR3-ANCA Vasculitis

“…Bands of interest were subjected to in-gel tryptic digest and analyzed by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS). 17 The molecular weights of the tryptic peptides from the larger protein band aligned with plasminogen, whereas those from the lower band aligned with ␤ 2 -glycoprotein-1. The middle protein band could not be positively identified.…”

Three isoforms of annexin I are preferentially expressed in normal esophageal epithelia but down-regulated in esophageal squamous cell carcinomas

“…Altered-expression protein spots from the preparative gels were digested in a modified four-step procedure as described by Beranova-Giorgianni and Desiderio (2000). Briefly, (1) protein spots were excised from the gels, washed in 50 mM NH 4 HCO 3 /acetonitrile (60/40) and dried by vacuum centrifugation; (2) 12 mg/ml sequencing grade modified porcine trypsin in 50 mM NH 4 HCO 3 digestion buffer was added to the dried gel pieces and incubated at 378C, 12 -16 h; (3) Peptides were recovered by three extractions of 30 ml of the digestion mixture with 50% ACN/5% TFA; (4) The recovered extracts were dried in a Speed-Vac and resuspended with 10 -15 ml 50% ACN/2.5% TFA.…”