Systematic Identification of the Subproteome of Escherichia coli Cell Envelope Reveals the Interaction Network of Membrane Proteins and Membrane-Associated Peripheral Proteins

Abstract

Membrane proteins of Gram-negative bacteria are key molecules that interface the cells with the environment. Despite recent proteomic identification of numerous oligomer proteins in the Escherichia coli cell envelope, the protein complex of E. coli membrane proteins and their peripherally associated proteins remain ill-defined. In the current study, we systematically analyze the subproteome of E. coli cell envelope enriched in sarcosine-insoluble fraction (SIF) and sarcosine-soluble fraction (SSF) by using proteomic methodologies. One hundred and four proteins out of 184 spots on 2D electrophoresis gels are identified, which includes 31 outer membrane proteins (OMPs). Importantly, our further proteomic studies reveal a number of previously unrecognized membrane-interacting protein complexes, such as the complex consisting of OmpW and fumarate reductase. This established complete proteomic profile of E. coli envelope also sheds new insight into the function(s) of E. coli outer envelope.

Quantitative Profile of the Uropathogenic Escherichia coli Outer Membrane Proteome during Growth in Human Urine

“…One study that pioneered the use of carbonate extraction and ASB-14 to efficiently solubilize and visualize the E. coli outer membrane identified 25 outer membrane proteins by using 2D-PAGE and mass spectrometry (23). More recently, sarcosine extraction was used to solubilize outer membrane proteins, including those that may be complexed and partition with inner membranes from E. coli, and the resulting proteomic analysis identified 31 outer membrane proteins (13). Here, for the first time, we define the profile of outer membrane proteins for an E. coli pathotype during growth in its natural environment.…”

Beta2-microglobulin is involved in the immune response of large yellow croaker to Aeromonas hydrophila: A proteomic based study

“…After equilibration for 15 min, the gel was transferred to the second dimension electrophoresis using 12% acrylamide gel. Gel spots containing the proteins of interest were manually excised from the Coomassie Blue stained gels and were in-gel digested with trypsin according to the procedure previously reported [15]. The resulting solution was analyzed on an autoflex MALDI-TOF/MS (BRUKER DAL-TONICS Company).…”

Functional characterisation of altered outer membrane proteins for tetracycline resistance in Escherichia coli

“…OM proteins were extracted as previously described [12]. In brief, C10 and T10 cultures were harvested at an optical density at 600 nm (OD 600 ) of 0.8 and disrupted by intermittent sonic oscillation.…”