Comparative analyses of proteins from XXXX biofilm and planktonic populations using metabolic labeling and mass spectrometry.

@entity3617 @entity2125 @entity1522 @entity2172 @entity2353 @entity844 @entity55 @entity10043 @entity327 @entity377 @entity6778 @entity7391
UNASSIGNED: BackgroundNon-typeable @entity327 (NTHi) is a nasopharyngeal commensal that can become an opportunistic pathogen causing infections such as @entity2172 , @entity55 , and @entity3617 . NTHi is known to form biofilms. Resistance of bacterial biofilms to clearance by host defense mechanisms and antibiotic treatments is well-established. In the current study, we used stable isotope labeling by @entity1522 in cell culture (SILAC) to compare the proteomic profiles of NTHi biofilm and planktonic organisms. Duplicate continuous-flow growth chambers containing defined media with either   light   (L) @entity6778 or   heavy   (H) 13C6-labeled @entity6778 were used to grow planktonic (L) and biofilm (H) samples, respectively. Bacteria were removed from the chambers, mixed based on weight, and protein extracts were generated. Liquid chromatography-mass spectrometry (LC-MS) was performed on the tryptic peptides and 814 unique proteins were identified with 99% confidence.ResultsComparisons of the NTHi biofilm to planktonic samples demonstrated that 127 proteins showed differential expression with p-values   0.05. Pathway analysis demonstrated that proteins involved in energy metabolism, protein synthesis, and @entity10043 , @entity7391 , @entity2353 , and @entity377 processes showed a general trend of downregulation in the biofilm compared to planktonic organisms. Conversely, proteins involved in transcription, DNA metabolism, and @entity844 and phospholipid metabolism showed a general trend of upregulation under biofilm conditions. Selected reaction monitoring (SRM)-MS was used to validate a subset of these proteins; among these were aerobic respiration control protein ArcA, @entity2125 nucleotidase and heme-binding protein A.ConclusionsThe present proteomic study indicates that the NTHi biofilm exists in a semi-dormant state with decreased energy metabolism and protein synthesis yet is still capable of managing oxidative stress and in acquiring necessary cofactors important for biofilm survival.
