Normal Urine Sugar Proteomics And Proteome Of E. Coli Dh5¦Á Secretion

As a necessary structure for membrane localization and secretion of mature proteins, N-linked glycosylation plays key roles in many biological processes. Changes of the glycosylation status and/or quantity of glycoproteins can be reflected in the profile changes of afffinity-enriched glycoproteins. In this project, N-linked glycoproteome from healthy human urine was studied because urine is a non-invasively accessible resource for biomarkers. Pooled urine samples from males and females were collected, and urine proteins were acetone precipitated. N-linked glycoproteins enriched with Concanavalin A affinity purification were separated and analyzed by SDS-PAGE-RPLC/MS/MS or 2D-LC/MS/MS. The tandem mass spectra of positive identifications were validated by AMASS software, and the peptide false positive rate was estimated using the reverse database searching method. A total of 225 urinary proteins were identified based on 2-hit criteria with reliability over 97％for each peptide. Among these proteins, 94 were identified in previous urine proteome works, 150 were annotated as glycoproteins or subunits of glycoproteins in SWISS-PROT, and 43 proteins were glycoproteins predicted by the software NetNGlyc 1.0 Server. Using the characteristic asparagine sequon detection, we found 48 possible N-glycosylation sites in 39 proteins. To enrich the details of urinary Con-A captured glycoprotein profile, 119 proteins identified by one hit were also listed. Compare to urine proteome, glycoproteome depletes some high abundant proteins and thus provides more detailed protein profile in addition to glycosylation information. It could be even a better choice over proteome itself in using urine sample to characterize human health status. The potential application of urinary N-glycoproteome in biomarker discovery is discussed. Secretory proteins from bacteria play key roles in many biological processes during their survival in external environment. With the development of proteome technology, it has been possible for the large scale detection of bacterial secretory proteins in one experiment. Here, secretory proteins of Escherichia coli DH5αcultured in M9 media were studied with one dimensional liquid chromatography-MS/MS. 53 proteins were obtained as positive identification, including enzymes, transport or binding proteins etc. which were mainly involved in adaptation to the bad surroundings, uptake, transport and metabolism of nutrients, as well as biosynthesis and metabolism of proteins, carbohydrates and lipids, et al. The causes that might lead to the release of these proteins into the media were discussed.