Isolation and characterization of hemin-permeable, envelope-defective mutants of Salmonella typhimurium

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Abstract
From S. typhimurium LT2 hemA (.delta.-aminolevulinic acid requiring) 15 mutants were isolated which grew on the hydrophobic compound hemin. All had increased sensitivity to antibiotics such as vancomycin, bacitracin, novobiocin, erythromycin, rifampin and oleandomycin and were considered to be envelope mutants (Env-). Half the mutants were rough, based on altered bacteriophage sensitivity and deoxycholate sensitivity; the remainder were smooth: 3 of the smooth mutants were studied in detail. They gave increased uptake of gentian violet but no increase in leakage of a periplasmic protein RNase I. The total membranes and fractions from sucrose gradient centrifugations representing inner and outer membranes of the wild type and 3 mutants were examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and isoelectric focusing-PAGE (IEF-PAGE). The major outer membrane proteins (MW 33,000, 34,000, 35,000 and 36,000) showed no or very little alterations in the Env- mutants. In SA1926 (env-52) 1 protein spot at MW 48,000, proven to be an outer membrane protein, was missing; a new spot appeared nearby and other proteins in this are of the gel were reduced. An Env+ transductant selected from this strain had the wild-type protein pattern restored. The 2 other Env mutants had similar but not identical changes in protein composition.