Two outer membrane proteins of Aeromonas hydrophila A6, isolated in a one- step affinity chromatography process based on carbohydrate reactivity, were found to be pore-forming molecules in artificial planar bilayer membranes. These carbohydrate-reactive outer membrane proteins (CROMPs; M(r)s, 40,000 and 43,000) were subjected to amino acid analysis. The amino acid profiles for these two outer membrane proteins were almost identical. A partial protein sequence of a 14-amino-acid fragment of the 40,000-Da protein revealed homology with outer membrane porins of Escherichia coli and A. hydrophila. CROMPs were compared with carbohydrate-reactive porins also extracted from outer membranes of A. hydrophila A6. These porins were isolated by using standard porin purification techniques (insolubility in 2% sodium dodecyl sulfate, solubility in 0.4 M NaCl, and Sephacryl S-200 gel filtration), and then Synsorb H type 2 affinity chromatography was done. The physical and functional properties of the carbohydrate-reactive porins and CROMPs were found to be identical. On the basis of pore-forming properties in planar lipid bilayers and channel inhibition with maltotriose solutions, a nonspecific, general diffusion porin and a LamB-like maltoporin were identified in both CROMP and carbohydrate-reactive porin preparations. To our knowledge, the use of carbohydrate reactivity to isolate channel-forming proteins from bacterial outer membranes has not been reported previously.
|Original language||English (US)|
|Number of pages||5|
|Journal||Infection and Immunity|
|State||Published - 1994|
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