The sensitivity of the outer and cytoplasmic membranes of Escherichia coli to detergent was examined by isopycnic sucrose density gradient centrifugation. Sodium lauryl sarcosinate (Sarkosyl) was found to disrupt the cytoplasmic membrane selectively under conditions in which Triton X-100 and dodecyl sodium sulfate solubilized all membrane protein. These results were verified by gel electrophoresis; membrane proteins solubilized by Sarkosyl were identical to those of the cytoplasmic membrane. The presence of Mg2+ during treatment with Sarkosyl was found to afford partial protection of the cytoplasmic membrane from dissolution.

From a lysogen with A integrated in the leu operon, specialized transducing phages that carry the cell division, murein biosynthesis, and envelope permeability genes located about 0.5 min to the right of leu were isolated. These phages were used to identify the previously undiscovered cell division gene sep. A genetic map proves that sep is located in the sequence leuA sep murE murF murC ddl ftsA envA. A physical map of this region was prepared by heteroduplex analysis of the phage DNAs. Overlapping segments of host DNA extended rightward for as much as 26.4 kilobase pairs from the prophage insertion point (thought to be in leuA) to include all the genes through envA. pi contained 0.02 M ethylenediaminetetraacetate (10 to 365 i), 1 N NaOH (5 il), Xinim4cIt6Sami7 phage stock (5 i1), and transducing phage stock (5 to 30 p1), added in that order. After 30 min at room temperature, J. BAcTroi.on August 1, 2020 by guest

The synthesis of membrane protein after infection with bacteriophage T4 was examined. Protein constituents of both the cytoplasmic and outer membrane are made during the infective cycle. In addition, newly synthesized membrane protein is found in material which has a buoyant density greater than that of either of the two host membrane fractions. Polyacrylamide gel analyses and solubilization studies using the detergent Sarkosyl indicate that synthesis of most of the membrane proteins made during the first 5 min of infection is directed by bacterial genes. New membrane proteins synthesized at times greater than 6 min after infection appear to be distinct from those of the host, and new proteins of the outer membrane are different from those of the inner. Proteins in the new dense membrane fraction are similar to those of the outer membrane.

Acidic proteases were extracted and purified from the stomach of orange roughy (Hoplostethus atlanticus). Protease I and II were glycoproteins with molecular weights of 33.5 and 34.5 KDa, respectively. Protease I had an isoelectric point of 5.30. The two forms of protease II (a and b) had isoelectric points of 4.35 and 4.40, respectively, and N‐terminal sequence identity for 12 amino acids. The proteases exhibited optimal temperature activity at 37C. They had high activity at low temperatures and low thermal stability compared to mammalian pepsins. They were stable in the pH range of 2–4.5 and unstable above pH 6.5. Protease I and II had pH optima of 2.5 and 3.5, respectively, and K m’values for the hydrolysis of hemoglobin (pH 3.0, 37C) of 124 μM and 517 μM, respectively. Enzyme activities were inhibited by pepstatin A and high NaCl concentrations, and were slightly stimulated by Ca2+ and Cu2+.

The "Mg2+-Sarkosyl crystals" (M band) technique distinguishes between membrane-bound and free intracellular DNA. This procedure was employed to investigate the nature of the reactions necessary to convert input T4 DNA to a rapidly sedimenting form. Energy poisoning inhibits this attachment reaction. Neither protein nor DNA synthesis appears to be required, but experiments with rifampin and extensively irradiated T4 suggest that RNA synthesis is involved. These results were confirmed by a second procedure for the determination of rapidly sedimenting DNA.