Purification and characterization of a lytic peptidase produced by Bdellovibrio bacteriovorus 6-5-S

Abstract

Bdellovibrio bacteriovorus 6-5-S releases at least two enzymes into the culture fluid when grown on autoclaved cells of Spirillum serpens VHL. One of the enzymes is bacteriolytic and the other is proteolytic. The lytic enzyme was purified by a factor of 3000 using ammonium sulfate fractionation, Sephadex G-100 gel filtration, and the anion exchanger DEAE-Sephadex. The lytic enzyme degrades peptidoglycan of S. serpens by hydrolyzing the diaminopimelic acid – alanine bond in the tetrapeptide chain. Ca2+ or Mg2+ exerted little or no effect on the activity of the lytic enzyme. The enzyme had a molecular weight of 40 000 as indicated by gel filtration. Crude preparations were unstable at 4C. The second enzyme, a protease that digested Azocoll, was purified by a factor of 7 by ammonium sulfate fractionation followed by gel filtration with Sephadex G-100. The protease was eluted in the void volume from a Sephadex G-100 column and therefore may have a molecular weight of at least 100 000. Its activity was enhanced by additions of Ca2+ and Mg2+. The enzyme was stable at 4C.