Isolation and Characterization of a New Extracellular Bacteriolytic Endopeptidase of Lysobacter sp. XL1

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O. A. Stepnaya^1*, I. M. Tsfasman¹, I. A. Logvina¹, L. P. Ryazanova¹, T. A. Muranova², and I. S. Kulaev¹

¹Skryabin Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, 142292 Pushchino, Moscow Region, Russia; fax: (7-095) 923-3602; E-mail: Stepnaya@ibpm.pushchino.ru

²Branch of Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russia; fax: (027) 790-527; E-mail: Muranova@fibkh.serpukhov.su

^* To whom correspondence should be addressed.

Received October 6, 2004; Revision received December 16, 2004

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The previously unstudied bacteriolytic enzyme L₄ was isolated from the culture liquid of the bacterium Lysobacter sp. XL1 in electrophoretically homogeneous state. The enzyme L₄ is a diaminopimelinoyl-alanine endopeptidase relative to peptidoglycan of Lysobacter sp. XL1. The enzyme is an alkaline protein of ~21 kD. The N-terminal amino acid sequence of the enzyme has been determined -- A V V N G V N Y V Gx T T A ... The maximal activity of the enzyme was observed in 0.05 M Tris-HCl at pH 8.0 and 50-55°C. The half-inactivation temperature of the enzyme is 52°C. The endopeptidase L₄ is not a metalloenzyme since it is not affected by EDTA. The enzyme is inhibited by p-chloromercuribenzoic acid by 72% and by phenylmethylsulfonyl fluoride by 43%, which indicates the involvement of serine and thiol groups in its functioning.

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KEY WORDS: bacteriolytic enzyme, Lysobacter sp., lysoamidase, purification, properties

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