Abstract
Acinetobacter baumannii, especially multidrug resistant Acinetobacter baumannii, is a notable source of pressure in the areas of public health and antibiotic development. To overcome this problem, attention has been focused on membrane proteins. Different digestion methods and extraction detergents were examined for membrane proteome sample preparation, and label-free quantitative and targeted proteome analyses of the polymyxin B-induced Acinetobacter baumannii ATCC 19606 membrane proteome were performed based on nano LC-MS/MS. Ultracentrifugation of proteins at a speed of 150,000×g, digestion by trypsin, filter-aided sample preparation, and detergents such as lauryldimethylamine-N-oxide were proved as a fast and effective way for identification of membrane proteome by nano LC-MS/MS. Upon treatment with polymyxin B, expression levels of 15 proteins related to membrane structure, transporters, cell surface, and periplasmic space were found to be significantly changed. Furthermore, targeted proteome was also used to confirm these changes. A relatively rapid membrane proteome preparation method was developed, and a more comprehensive view of changes in the Acinetobacter baumannii membrane proteome under polymyxin B pressure was obtained.
Highlights
Acinetobacter baumannii (A. baumannii) is a Gram-negative bacterium that can cause serious nosocomial infection (Lin and Lan, 2014) and has developed extensive antimicrobial resistance with a high mortality rate (Vazquez-Lopez et al, 2020)
The World Health Organization (WHO) reported that A. baumannii was one of the most serious ESKAPE organisms (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, A. baumannii, Pseudomonas aeruginosa, and Enterobacter cloacae) which could effectively escape the actions of antibacterial drugs (Boucher et al, 2009)
We evaluated the effectiveness of different digestion enzymes, digestion conditions, and detergents [ionic, nonionic, and zwitterionic detergents (lauryldimethylamine-N-oxide, LDAO or 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate, CHAPS)] in A. baumannii membrane proteome analysis by nano LC-high resolution MS and developed an effective and rapid workflow for A. baumannii membrane proteome analysis
Summary
Acinetobacter baumannii (A. baumannii) is a Gram-negative bacterium that can cause serious nosocomial infection (Lin and Lan, 2014) and has developed extensive antimicrobial resistance with a high mortality rate (Vazquez-Lopez et al, 2020). With the increase in multidrug resistance (MDR), A. baumannii showed resistance to many first-line antibiotics, even colistin and polymyxin B (Olaitan et al, 2014). Membrane proteins play vital roles in defense and immunity and in essential physiological functions, such as the generation or conversion of energy, the import or export of metabolites, the extrusion of toxic substances or antibiotics, and the homeostasis of metal ions (Ansgar and Dirk, 2010).
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