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Calibration was updated before each acquisition using a standard peptide mixture according to instrument protocol.
Prior to analysis the MS method was calibrated using a standard peptide mix (Bruker Daltonics) and the matrix cluster peak from CHCA at m/z 379.1 Da.
All mass spectra were externally calibrated using a standard peptide mixture containing des-Arg-Bradykinin (904.4681), angiotensin I (1296.6853), 1 17 (2093.0867) and 18 39 (2465.1989) adrenocorticotropic hormon fragments.
Dantuma et al constructed a fusion of GFP to Ubiquitin (Ubi[G76V]-GFP) using a standard peptide bond at the N-terminus [21], Another proteasome sensor construct, which is a GFP fusion to an artificial peptide, CL1, identified in yeast has been designed by Bence et al ([22].
Calibration was carried out using a standard peptide mixture.
The instrument was calibrated using a standard peptide mixture (Bruker Daltonic GmbH).
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The single-chain dimeric peptides were synthesized and produced in 10-mg amounts (AMS Biotechnology, Oxon, England) using a standard solid-phase peptide synthesis on Wang resin with fluorenylmethoxycarbonyl (Fmoc) -protected amino acids.
External mass calibration of the hybrid mass spectrometer is performed using a standard calibration mixture of different peptides and matrix components.
The peptidic spacers H-l-Phe-l-Phe-l-Lys(Boc -OH (FFK(Boc -OH) and H-d-Phe-d-Phe-d-Lys(Boc)-OH (FFK(Boc), 6) were synthesized via solid phase peptide synthesis using a standard Fmoc protocol.
The exact number of moles for each peptide was calculated using a standard curve comparing the concentration and area.
Peptides were synthesized using a standard fluorenylmethyl (Fmoc) procedure on solid support, as previously described.
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