ABSTRACT
Interferon stimulated gene 15 kDa protein (ISG15) is the first ubiquitin-like protein identified, which plays vital roles in a variety of fields including viral infection and immunological regulation. In this study, liquid chromatography-tandem mass spectrometry was used to analyze ISG15-modified proteins in A549 cells in response to infection by influenza virus, which was enriched by immunoprecipitation. A total of 22 cellular host proteins were identified in A549 cells infected by influenza virus, including ubiquitin-like ISG15 protein, cyclin-T1, heat shock protein 71 kDa, caldesmon, eukaryotic translation initiation factor, and so on. Besides, non-structural protein (NS1) from influenza virus was also identified. Among the 22 host proteins identified, 6 proteins were also identified in the control non-infected A549 cells, including annexin A1, fructose-bisphosphate aldolase A, ATP synthase subunit g, enolase, actin, and tubulin. Bioinformatics analysis revealed that the identified ISG15-modified host proteins induced by influenza virus infection could be classified into 9 protein classes: chaperone, oxidoreductase, enzyme modulator, transferase, nucleic acid binding, transcription factor, kinase, cytoskeletal protein, and structural protein. This study provided a specific and effective tool for analyzing ISG15-modified proteins in proteome level.
ABSTRACT
Abstract Formaldehyde has been widely employed to immobilize clinical tissue specimens, inactivate toxins and viruses in biomedical fields. Formaldehyde can react with active groups in bio-molecules such as proteins, resulting in protein cross-linking, inactivation, and immobilization. By using several standard peptides and tryptic peptides from matrix protein of influenza virus as experimental models, we studied the chemical modifications of peptides and proteins with formaldehyde by matrix-assisted laser desorption ionization time-of-flight mass spectrometry and nano-electrospray quadruple time-of-flight tandem mass spectrometry. The reaction between formaldehyde and peptides was performed under the same conditions as those during inactivation of virus (4℃, 0. 025% Formalin (V/V), 37% formaldehyde solution (w/w), and 72 h). The results indicated that under above conditions, formaldehyde could react with amino group of N-terminus of standard peptide to generate a methylol adduct, which was further condensed into an imine to generate+12 Da product. Besides, formaldehyde could react with side chain of two amino acids such as arginine and lysine, yielding +12 Da product respectively. The analysis of the reaction between formaldehyde and tryptic peptides from matrix protein of influenza virus showed that +24 Da products could be detected in most peptides due to combinational contribution from N-terminus of peptide (+12 Da ) and side chain of C-terminal arginine or lysine (+12 Da) . Moreover, a +36 Da product was detected for a peptide with miss-cut site. The results indicated that low-concentration formaldehyde primarily reacted with amino group on N-termini of peptides and proteins, as well as the side chains of arginine and lysine residues. The present study suggested an effective mass spectrometry-based method for analyzing the reaction between low-concentration formaldehyde and peptides and proteins, thus provided strategies for interpretation for the mass spectra of reaction products.
ABSTRACT
Objective To analysis the risk factors and drug resistance of multidrug-resistant pseudomonas aeruginosa(MDRP) infection and to provide the basis for clinical anti-infective therapy .Methods Predisposing factors and drug resistance to clinical commonly used antibacterial drugs of MDRP which were separated from September 2010 to December 2011 were adopted for retro-spective analysis .Results A total of 235 hospital MDRP infection were collected ,97 multidrug resistant strains were concluded ,the separation rate was 41 .2% .The separation rate of ICU and neurosurgery were higher ,account for 35 .54% and 22 .31% respective-ly .The main resource of specimen is respiratory tract ,about 75 .21% .The resistance of MDRP was very serious .The resistant rate of Amikacin was the lowest ,account for 37 .11% ,followed by piperacillin/tazobactam ,ceftazidime ,account for 47 .4% and 48 .45%respectively .The resistant rate of other antimicrobial agents were greater than 50% .Advanced age ,serious underlying diseases ,long hospital stay ,long-term repeated application of broad-spectrum antimicrobial drugs ,admission to ICU ,invasive treatment operations were the risk factors for MDRP resistance .Conclusion The resistance is very serious ,in order to reduce the generation and spread of drug-resistant strains ,the hospital infection control ,drug resistance monitoring ,rational use of antimicrobial drugs and prevention of cross-infections should be strengthened .