Refractile Granular Inclusions in a Temperature-sensitive Mutant of Escherichia coli K-12

At the restrictive temperature, cells of a temperature-sensitive mutant of E. coli K-12 lyse, and the culture medium is seen to contain ghosts of cells containing large inclusions that are refractile when viewed by phase-contrast microscopy. The addition of sodium chloride (l %) or sucrose (12, 5 %) protects the cells from lysis and under these conditions the refractile inclusions are seen in the intact cells, which are short filaments. At elevated temperatures cells become sensitive to a wide range of inhibitors. Isolated inclusions consist almost entirely of protein and are digested by Pronase or trypsin. Electrophoresis of inclusions on sodium dodecyl sulphate-polyacrylamide gels shows them to contain a number of protein components. Ultrastructural studies show that granules are commonly found close to the internal surface of the cytoplasmic membrane. In section they are seen to be grainy in appearance but have no ordered structure. Dissociation of the granules by sodium deoxy-cholate or trypsin leads to the formation of spirally wound filaments with a subunit structure.