Evaluation of SDS‐polyacrylamide gel systems for the study of outer membrane protein profiles of clinical strains of Acinetobacter baumannii

Abstract

The outer membrane protein (OMP) profiles of 23 blood isolates of Acinetobacter baumannii representing all the different antimicrobial susceptibility patterns observed during a 3-year period in a Spanish hospital were studied. OMPs extracted from envelopes of sonicated cells after solubilisation with 2% of N-lauryl-sarcosinate were analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) using the Laemmli's buffers. Eight running gel systems differing in the concentration of polyacrylamide (8%, 10% and 12%) and in the absence or presence of urea (4 M and 6 M) were used in a preliminary study analysing the OMP profiles of four clonally unrelated strains of A. baumannii. When this study was completed, the OMPs of the 23 A. baumannii were analysed in 10% SDS-polyacrylamide gels with 6 M urea and in 12% SDS-polyacrylamide gels. Ten OMP profiles were observed in 10% SDS-polyacrylamide gels with 6 M urea, whereas only 5 OMP profiles were visualised using 12% SDS-polyacrylamide gels. The OMP profiles obtained in 10% SDS-polyacrylamide gels with 6 M urea only partially correlated with those observed in 12% SDS-polyacrylamide gels. In conclusion, the use of 10% SDS-polyacrylamide gels with 6 M urea is recommended for the study of OMP profiles of A. baumannii.

Platelet-activating Factor Receptor Initiates Contact of Acinetobacter baumannii Expressing Phosphorylcholine with Host Cells

“…baumannii clinical isolate 77wt, collected in Spain in 2000 (27), its derivative ciprofloxacin-resistant strain 77R, and six other A. baumannii clinical strains were used (28,29). ATCC 17978wt and its derivative porinD mutant were also used (supplemental Table S1) (30).…”

“…ChoP Immunoblotting-Outer membrane fractions from ATCC 17978 wt, PSAB03, 77wt, 77R, and six other A. baumannii clinical strains were isolated as described previously (29). Western blot was performed using 10% SDS-PAGE with antiChoP TEPC-15 mAb as primary antibody and horseradish peroxidase-conjugated goat anti-mouse IgG antibody as secondary antibody.…”

Loss of the 29-Kilodalton Outer Membrane Protein in the Presence of OXA-51–Like Enzymes inAcinetobacter baumanniiIs Associated with Decreased Imipenem Susceptibility

“…The pellet was washed once with 2% sodium N-lauryl sarcosinate, collected again as described previously, and finally resuspended in 20 mM Tris-HCl (pH 8.0)-0.1 mM ethylenediaminetetraacetic acid -1% sodium dodecyl sulfate (SDS) for further analyses. 13,28 SDS-polyacrylamide gel electrophoresis and Coomassie blue staining. The OMP prepared as above was analyzed by SDS-polyacrylamide gel electrophoresis (PAGE) with 12% polyacrylamide gels.…”

Safety Assessment of Transgenic Organisms, Volume 4

“…They contribute to the efficiency of the bacteria to utilize iron, a host resources that is limiting, and therefore contribute to the virulence of a particular strain (Dorsey et al, 2003). SDS-polyacrylamide gel systems have been used for the study of outer membrane protein profiles of clinical strains of A. baumannii (Cuenca et al, 2003) The outer membrane of non-clinical Acinetobacter strains also contains proteins of great importance for the interaction of the bacteria with its environment. Membrane proteome of A. radioresistens S13 during aromatic exposure has been studied with two-dimensional gel electrophoresis.…”

“…They contribute to the efficiency of the bacteria to utilize iron, a host resources that is limiting, and therefore contribute to the virulence of a particular strain (Dorsey et al, 2003). SDS-polyacrylamide gel systems have been used for the study of outer membrane protein profiles of clinical strains of A. baumannii (Cuenca et al, 2003) 160 -PART 2. DOCUMENTS ON MICRO-ORGANISMS SAFETY ASSESSMENT OF TRANSGENIC ORGANISMS: OECD CONSENSUS DOCUMENTS: VOLUME 4 © OECD 2010…”