microbiology, Vol 140, 797-805, Copyright © 1994 by Society for General Microbiology

ARTICLES

Isolation and characterization of the outer-membrane proteins of Burkholderia (Pseudomonas) pseudomallei

N Gotoh, NJ White, W Chaowagul and DE Woods
Department of Microbiology and Infectious Diseases, University of Calgary Health Sciences Centre, Alberta, Canada.

Membranes obtained from whole-cell lysates of Burkholderia (Pseudomonas) pseudomallei (strain 319a) were separated into four fractions by sucrose density gradient centrifugation. Membranes were characterized by enzymic and chemical analyses, and by SDS-PAGE. Cytoplasmic membranes and two forms of outer membranes (OM-1, OM-2) were detected. The major outer-membrane proteins had M(r) values of 70,000, 38,000, 31,000, 24,000 and 17,000. To determine which outer- membrane proteins were common to B. pseudomallei strains, OM-1 fractions from 12 different strains were prepared. SDS-PAGE analysis of these fractions demonstrated that the five major outer-membrane proteins were common to the strains tested. Further studies have shown that an M(r) 110,000 protein, which is oligomeric in that it migrates as an M(r) 38,000 protein upon heating at 95 degrees C and which is peptidoglycan-associated, serves as a porin in B. pseudomallei. Using proteoliposomes reconstituted from this protein and phospholipid, it was demonstrated by the liposome-swelling assay that this protein acts as a porin through which small saccharides may diffuse. Further characterization of this M(r) 38,000 protein will be important in delineating the role of this molecule in the permeability of the B. pseudomallei outer membrane.

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