Bacillus species LU4 Is an Effective Producer of Thermostable Site-Specific Endonuclease BspLU4I, an Isoschizomer of AvaI

I. V. Svadbina¹, E. N. Zheleznyakova², L. A. Zheleznaya¹, and N. I. Matvienko^2*

¹Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences, Pushchino, Moscow Region 142290, Russia; fax: (0967) 790-553

²Institute of Protein Research, Russian Academy of Sciences, Pushchino, Moscow Region 142290, Russia; fax: (095) 924-0493; E-mail: nikmatv@vega.protres.ru

^* To whom correspondence should be addressed.

Received March 6, 2002; Revision received May 22, 2002

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The site-specific endonuclease BspLU4I was discovered in the thermophilic Bacillus species LU4 strain and purified to functionally pure state by chromatography on blue agarose, hydroxyapatite HTP, and heparin-Sepharose columns. Analysis of cleavage patterns of different DNAs with known nucleotide sequences demonstrated that the enzyme recognizes the CPyCGPuG site on the DNA. Cleavage points in the sequence were determined with the elongated primer method. It was shown that the endonuclease is an isoschizomer of AvaI. The final yield of the enzyme is 2.25·10⁶ units per g wet biomass.

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KEY WORDS: restriction endonucleases, isoschizomers, AvaI, DNA size markers

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