Lytic Enzyme from Lysates of Streptomyces venezuelae Infected with Actinophage MSP2

Abstract

A lytic enzyme was purified 600-fold with 12% recovery from lysates of Streptomyces venezuelae S13 infected with actinophage MSP2. The purified enzyme preparation was homogeneous as shown by polyacrylamide electrophoresis. The enzyme was active over a p H range 6.0 to 9.0 with a maximum at p H 7.5. The p H profile for stability was sharp, with an optimum at p H 7.5. Maximal activity occurred between 30 and 35 C. The enzyme was stable at 20 C or less. A 30-min exposure to 25, 30, 35, 40, 45, and 50 C produced an inactivation of 3, 40, 77, 82, 93, and 100%, respectively. Lytic activity was stimulated fivefold by either 5 × 10 −3 m Mg 2+ or Mn 2+ and three- and twofold by Ca 2+ and Ba 2+ , respectively. Addition of Na + , K + , NH 4 + , or Li + to the tris(hydroxymethyl)aminomethane-hydrochloride buffer did not alter the rate of lysis. Enzyme activity was inhibited 74 and 27% by 10 −4 and 10 −5 m ethylenediaminetetraacetic acid (EDTA), respectively. The inhibition by EDTA was reversed partially by addition of Mg 2+ . Lytic activity was abolished by either 5 × 10 −4 m HgCl 2 or p -hydroxymercuribenzoate, whereas 5 × 10 −4 m CuSO 4 inhibited 72%. Cell wall solubilization paralleled the release of N -terminal amino groups and reached a level of 0.23 μmole per mg of cell walls. No release of reducing power was detected in treated or untreated cell wall suspensions. Tests for proteolytic activity were negative.