Abstract
BackgroundIt is widely believed that integral outer membrane (OM) proteins in bacteria are able to diffuse laterally in the OM. However, stable, immobile proteins have been identified in the OM of Escherichia coli. In explaining the observations, a hypothesized interaction of the immobilized OM proteins with the underlying peptidoglycan (PG) cell wall played a prominent role.ResultsOmpA is an abundant outer membrane protein in E. coli containing a PG-binding domain. We use FRAP to investigate whether OmpA is able to diffuse laterally over long-range (> ~100 nm) distances in the OM. First, we show that OmpA, containing a PG binding domain, does not exhibit long-range lateral diffusion in the OM. Then, to test whether PG interaction was required for this immobilization, we genetically removed the PG binding domain and repeated the FRAP experiment. To our surprise, this did not increase the mobility of the protein in the OM.ConclusionsOmpA exhibits an absence of long-range (> ~100 nm) diffusion in the OM that is not caused by its PG binding domain. Therefore, other mechanisms are needed to explain this observation, such as the presence of physical barriers in the OM, or strong interactions with other elements in the cell envelope.
Highlights
It is widely believed that integral outer membrane (OM) proteins in bacteria are able to diffuse laterally in the OM
We find that full-length OmpA exhibits an absence of long-range (> 100 nm) diffusion in the OM
Since its discovery as fluorescent periplasmic reporter in E. coli [19], mRFP1/mCherry [18] has been used in several fusion constructs that sub-localize to the PG/OM layer, without interfering with their function
Summary
It is widely believed that integral outer membrane (OM) proteins in bacteria are able to diffuse laterally in the OM. Little information exists on the mobility of (integral) outer membrane proteins (OMPs) in the bacterial OM. Experimental evidence suggests that integral outer membrane protein IcsA is able to diffuse laterally over micron-ranges in the OM [3]. Recent work on the mobility of integral OMP LamB suggests that it is confined to a region of size ~50 nm [4,5]. This was based on the motion of a
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