Abundant ions corresponding to the gas-phase cleavage of the Asp-Pro and Asp-Xxx bonds of peptides in the process of matrix-assisted laser desorption were observed using a time-of-flight mass spectrometer equipped with both linear and reflector mass analyzers. Peptides containing the N-terminal sequence, Asp-Pro ... from an endoproteinase Asp-N digest yielded one peak in the molecular ion region in the linear mode and two equally abundant peaks in the reflector mode TOF mass spectra. The lower molecular masses in the reflector mode mass spectra could be eliminated by removing the Asp residue or derivatizing its side-chain carboxyl group. The observed mass differences did not correspond to any amino acid; however, by lowering the potential of the reflector to correct for the energy loss the mass difference was determined to be 115 Da, i.e., Asp. The extent and rate of this decomposition was compared with that obtained using a four-sector tandem mass spectrometer in the MS/MS mode of operation without and with a collision gas at collision cell potentials of 3.0 and 9.86 kV. These data suggest the Asp-Pro peptide bond is more labile than other peptide bonds in the gas phase. Abundant metastable decomposition of internal Asp-Pro bonds was also observed in larger peptides and proteins. Based on these latter data, a mechanism for this gas-phase cleavage is proposed.(ABSTRACT TRUNCATED AT 250 WORDS)

Peptide sequencing via tandem mass spectrometry (MS/MS) is one of the most powerful tools in proteomics for identifying proteins. Because complete genome sequences are accumulating rapidly, the recent trend in interpretation of MS/MS spectra has been database search. However, de novo MS/MS spectral interpretation remains an open problem typically involving manual interpretation by expert mass spectrometrists. We have developed a new algorithm, SHERENGA, for de novo interpretation that automatically learns fragment ion types and intensity thresholds from a collection of test spectra generated from any type of mass spectrometer. The test data are used to construct optimal path scoring in the graph representations of MS/MS spectra. A ranked list of high scoring paths corresponds to potential peptide sequences. SHERENGA is most useful for interpreting sequences of peptides resulting from unknown proteins and for validating the results of database search algorithms in fully automated, high-throughput peptide sequencing.

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The utility of matrix-assisted laser desorption time-of-flight (MALD-TOF) mass spectrometry for the analysis of recombinant glycopeptides is discussed and compared to information which may be obtained by fast atom bombardment mass spectrometry (FABMS) and tandem mass spectrometry (MS/MS). MALD-TOF appears to be 10-100 times more sensitive than FAB MS for the analysis of underivatized glycopeptides, providing qualitative site-specific information regarding the carbohydrate microheterogeneity without the extensive isolation and derivatization procedures required to obtain similar information by FAB MS. Analysis of a digest mixture in the positive and negative ion mode of MALD-TOF indicated that, in mixtures, sialylated glycopeptides are preferentially detected in the negative ion mode. The determination of the molecular masses of a glycopeptide with MALD-TOF prior to and after treatment with a variety of specific glycosidases, often without removal of the buffers, coupled to a comparison of molecular mass information available from a carbohydrate database facilitates the assignment of a carbohydrate composition. The vast majority of the molecular ion signal observed in the linear mode for sialylated glycopeptides are metastable ions. Reflector mass spectra reveal a shift to lower mass consistent with the loss of most of the neuraminic acid residues. The loss of Hex and HexNAc residues is also observed. Sequential lowering of the reflector potential reveals structurally significant fragment ions representing the carbohydrate and peptide portions of the molecule.

Objective: Evaluate the safety and tolerability of beloranib, a fumagillin-class methionine aminopetidase-2 (MetAP2) inhibitor, in obese women over 4 weeks. Design and Methods: Thirty-one obese (mean BMI 38 kg/m 2 ) women were randomized to intravenous 0.1, 0.3, or 0.9 mg/m 2 beloranib or placebo twice weekly for 4 weeks (N ¼ 7, 6, 9, and 9). Results:The most frequent AEs were headache, infusion site injury, nausea, and diarrhea. Nausea and infusion site injury occurred more with beloranib than placebo. The most common reason for discontinuation was loss of venous access. There were no clinically significant abnormal laboratory findings. In subjects completing 4 weeks, median weight loss with 0.9 mg/m 2 beloranib was À3.8 kg (95% CI À5.1, À0.9; N ¼ 8) versus À0.6 kg with placebo (À4.5, À0.1; N ¼ 6). Weight change for 0.1 and 0.3 mg/m 2 beloranib was similar to placebo. Beloranib (0.9 mg/m 2 ) was associated with a significant 42 and 18% reduction in triglycerides and LDL-cholesterol, as well as improvement in C-reactive protein and reduced sense of hunger. Changes in b-hydroxybutyrate, adiponectin, leptin, and fibroblast growth factor-21 were consistent with the putative mechanism of MetAP2 inhibition. Glucose and blood pressure were unchanged. Conclusions: Beloranib treatment was well tolerated and associated with rapid weight loss and improvements in lipids, C-reactive protein, and adiponectin.