Isolation and Structural Characterization of a Cytotoxic L-Amino Acid Oxidase from Agkistrodon contortrix laticinctus Snake Venom: Preliminary Crystallographic Data

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• 作者：Souza ; Dulce H. F. ; Eugenio ; Luiz M. ; Fletcher ; Jeffrey E. ; Jiang ; Ming-Shi ; Garratt ; Richard C. ; et. al.
• 刊名：Archives of Biochemistry and Biophysics
• 出版年：1999
• 出版时间：August 15, 1999
• 年：1999
• 卷：368
• 期：2
• 页码：285-290
• 全文大小：436 K

文摘

We have purified a cytotoxic L-amino acid oxidase (LAO) from Agkistrodon contortrix laticinctus snake venom by means of Superdex-200 gel filtration, followed by phenyl-Sepharose CL-4B chromatography. The purified enzyme (ACL LAO) is a dimer on gel filtration, with a M_r of 60,000 for the monomer as estimated by SDS–PAGE. LAO activity was tested against 15 amino acids, but only 9 were oxidized by the enzyme, suggesting that it presents some degree of specificity. ACL LAO has apoptosis-inducing activity in an HL-60 cell culture assay. After 24 h treatment with 25 μg/ml of ACL LAO, the typical DNA fragmentation pattern of apoptotic cells was observed on agarose gel electrophoresis. NMR analysis showed the presence of a flavin mononucleotide prosthetic group. To solve its 3-D structure, crystals of the purified protein were grown in 0.1 M Tris–HCl, pH 8.5, and 2 M (NH₄)₂SO₄. Diffraction data collected to 3.5 Å showed that the protein crystallized in the tetragonal system, with unit cell a = b = 103.22 Å, c = 183.45 Å. This is the first report of preliminary crystallization data for a snake venom L-amino acid oxidase.