“…The snR30 and snR42 snoRNP particles were isolated from 100 L of whole-cell yeast extract as described above+ The fractions corresponding to snR30 and snR42 snoRNPs were then phenol/chloroform extracted and the proteins precipitated with acetone+ The resulting proteins were separated on a 13% high-TEMED SDS polyacrylamide gel and stained with colloidal Coomassie blue (Sigma)+ The protein bands were excised from the gel, washed, reduced and alkylated, and finally digested with trypsin as previously described (Shevchenko et al+, 1996)+ After 3 h digestion, 0+3 mL of the supernatant were taken out and used for MALDI peptide mass mapping+ For nanoelectrospray mass spectrometry, the peptides were extracted after digestion over night and desalted into the spraying capillary of the nanoelectrospray source, as described (Shevchenko et al+, 1996;+ Mass spectra were acquired on Bruker REFLEX Matrix Assisted Laser Desorption Ionization time-of-flight mass spectrometer (Bruker-Franzen, Bremen, Germany) equipped with delayed extraction ion source+ The mixture of saturated solution of alpha-cyano hydroxycynnamic acid in acetone and nitrocellulose was used as a thin layer matrix (Vorm et al+, 1994;Jensen et al+, 1996)+ The 0+3 mL of digestion supernatant was deposited into an acidified water droplet on the surface of the matrix (Jensen et al+, 1996)+ Trypsin autolysis products were used for internal calibration that give average accuracy better than 50 ppm+ For identification of proteins using the sequence tag approach (Mann & Wilm, 1994), tandem mass spectra were acquired on an API III triple quadrupole mass spectrometer (Perkin Ellmer-Sciex, Ontario, Canada) equipped with an updated collision cell and a nanoelectrospray source + The peptide ions were sequentially fragmented and from the fragment spectra partial sequence data was obtained for database searching (Mann & Wilm, 1994)+ All proteins were identified using PeptideSearch in nrdb (nonredundant database), which currently holds more than 250,000 entries+…”