Lysine-49-phospholipases A2 from Trimeresurus flavoviridis venom are membrane-acting enzymes

Yasuyuki Shimohigashi, Ayako Tani, Hiroshi Matsumoto, Kinichi Nakashima, Yoko Yamuguchi, Naoko Oda, Yukio Takano, Hiro o. Kamiya, Junji Kishino, Hitoshi Arita, Motonori Ohno

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Abstract

Basic proteins I and II (BP-I and BP-II) isolated from the venom of Trimeresurus flavoviridis (Habu snake) are isozymes of highly active Asp-49-phospholipase A2 (Asp-49-PLA2) and classified into the group Lys-49-PLA2. BP-II was found to elicit a strong contraction of Guinea pig ileum, and this activity was inhibited completely by 1 μM indomethacin, an inhibitor of the arachidonate cascade. BP-II was inactive in the Ca2+ -free medium, and p-bromophenacy lated His-48-BP-II was also inactive. BP-II exhibited no binding affinity for the cells expressinng PLA2 receptors. These results indicated that the contraction elicited by BP-II is due to the hydrolytic action of BP-II, liberating archidonic acid from the ileum phospholipid biomembranes. In spite of its limited lipolytic activities (av. 0.9% of Asp-49-PLA2) against monomers and micelles of synthetic phospholipids, BP-II hydrolyzed considerably strongly the phospholipids in the artificial bilayer vesicles. Arachidonic acid released from liposomes of β-arachidonoyl-γ-stearoyl-l-α-phosphatidylcholic was determined by HPLC, and the activity of BP-II was estimated to be about 75% as compared to Asp--49-PLA2. Liposomes encapsulating carboxyflurescein exhibited a strong dye-leakage induced by BP-II in a concentration-dependent manner, only in the Ca2+-containing buffer. The net result from all these observations was that BP-II, a Lys-49-PLA2, is an enzyme that hyrolyzes the membrane phospholipids. In contrast to BP-II, BP-I was found to be considerably weak in hydrolyzing membrane phospholipids, although its activities were distinct. BP-I and BP-Ii share a common sequence with the sole exception of Asp-67 (BP-I) and Asn-67 (BP-II) in the aligned sequences. This implies that the amino acid at position 67 of Lys-49-PLA2s is the residue required for discriminatory recognition of β-arachidonoyl-phospholipid membranes.

Original languageEnglish
Pages (from-to)1037-1044
Number of pages8
JournalJournal of biochemistry
Volume118
Issue number5
DOIs
Publication statusPublished - Oct 1 1995

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Phospholipases A2
Venoms
Lysine
Phospholipids
Membranes
Enzymes
Ileum
Liposomes
Trimeresurus
Snakes
Micelles
Arachidonic Acid
Indomethacin
Isoenzymes
Trimeresurus venoms
Buffers
Guinea Pigs
Coloring Agents
Monomers
High Pressure Liquid Chromatography

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

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Lysine-49-phospholipases A2 from Trimeresurus flavoviridis venom are membrane-acting enzymes. / Shimohigashi, Yasuyuki; Tani, Ayako; Matsumoto, Hiroshi; Nakashima, Kinichi; Yamuguchi, Yoko; Oda, Naoko; Takano, Yukio; Kamiya, Hiro o.; Kishino, Junji; Arita, Hitoshi; Ohno, Motonori.

In: Journal of biochemistry, Vol. 118, No. 5, 01.10.1995, p. 1037-1044.

Research output: Contribution to journalArticle

Shimohigashi, Y, Tani, A, Matsumoto, H, Nakashima, K, Yamuguchi, Y, Oda, N, Takano, Y, Kamiya, HO, Kishino, J, Arita, H & Ohno, M 1995, 'Lysine-49-phospholipases A2 from Trimeresurus flavoviridis venom are membrane-acting enzymes', Journal of biochemistry, vol. 118, no. 5, pp. 1037-1044. https://doi.org/10.1093/jb/118.5.1037
Shimohigashi, Yasuyuki ; Tani, Ayako ; Matsumoto, Hiroshi ; Nakashima, Kinichi ; Yamuguchi, Yoko ; Oda, Naoko ; Takano, Yukio ; Kamiya, Hiro o. ; Kishino, Junji ; Arita, Hitoshi ; Ohno, Motonori. / Lysine-49-phospholipases A2 from Trimeresurus flavoviridis venom are membrane-acting enzymes. In: Journal of biochemistry. 1995 ; Vol. 118, No. 5. pp. 1037-1044.
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abstract = "Basic proteins I and II (BP-I and BP-II) isolated from the venom of Trimeresurus flavoviridis (Habu snake) are isozymes of highly active Asp-49-phospholipase A2 (Asp-49-PLA2) and classified into the group Lys-49-PLA2. BP-II was found to elicit a strong contraction of Guinea pig ileum, and this activity was inhibited completely by 1 μM indomethacin, an inhibitor of the arachidonate cascade. BP-II was inactive in the Ca2+ -free medium, and p-bromophenacy lated His-48-BP-II was also inactive. BP-II exhibited no binding affinity for the cells expressinng PLA2 receptors. These results indicated that the contraction elicited by BP-II is due to the hydrolytic action of BP-II, liberating archidonic acid from the ileum phospholipid biomembranes. In spite of its limited lipolytic activities (av. 0.9{\%} of Asp-49-PLA2) against monomers and micelles of synthetic phospholipids, BP-II hydrolyzed considerably strongly the phospholipids in the artificial bilayer vesicles. Arachidonic acid released from liposomes of β-arachidonoyl-γ-stearoyl-l-α-phosphatidylcholic was determined by HPLC, and the activity of BP-II was estimated to be about 75{\%} as compared to Asp--49-PLA2. Liposomes encapsulating carboxyflurescein exhibited a strong dye-leakage induced by BP-II in a concentration-dependent manner, only in the Ca2+-containing buffer. The net result from all these observations was that BP-II, a Lys-49-PLA2, is an enzyme that hyrolyzes the membrane phospholipids. In contrast to BP-II, BP-I was found to be considerably weak in hydrolyzing membrane phospholipids, although its activities were distinct. BP-I and BP-Ii share a common sequence with the sole exception of Asp-67 (BP-I) and Asn-67 (BP-II) in the aligned sequences. This implies that the amino acid at position 67 of Lys-49-PLA2s is the residue required for discriminatory recognition of β-arachidonoyl-phospholipid membranes.",
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AU - Shimohigashi, Yasuyuki

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AU - Nakashima, Kinichi

AU - Yamuguchi, Yoko

AU - Oda, Naoko

AU - Takano, Yukio

AU - Kamiya, Hiro o.

AU - Kishino, Junji

AU - Arita, Hitoshi

AU - Ohno, Motonori

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N2 - Basic proteins I and II (BP-I and BP-II) isolated from the venom of Trimeresurus flavoviridis (Habu snake) are isozymes of highly active Asp-49-phospholipase A2 (Asp-49-PLA2) and classified into the group Lys-49-PLA2. BP-II was found to elicit a strong contraction of Guinea pig ileum, and this activity was inhibited completely by 1 μM indomethacin, an inhibitor of the arachidonate cascade. BP-II was inactive in the Ca2+ -free medium, and p-bromophenacy lated His-48-BP-II was also inactive. BP-II exhibited no binding affinity for the cells expressinng PLA2 receptors. These results indicated that the contraction elicited by BP-II is due to the hydrolytic action of BP-II, liberating archidonic acid from the ileum phospholipid biomembranes. In spite of its limited lipolytic activities (av. 0.9% of Asp-49-PLA2) against monomers and micelles of synthetic phospholipids, BP-II hydrolyzed considerably strongly the phospholipids in the artificial bilayer vesicles. Arachidonic acid released from liposomes of β-arachidonoyl-γ-stearoyl-l-α-phosphatidylcholic was determined by HPLC, and the activity of BP-II was estimated to be about 75% as compared to Asp--49-PLA2. Liposomes encapsulating carboxyflurescein exhibited a strong dye-leakage induced by BP-II in a concentration-dependent manner, only in the Ca2+-containing buffer. The net result from all these observations was that BP-II, a Lys-49-PLA2, is an enzyme that hyrolyzes the membrane phospholipids. In contrast to BP-II, BP-I was found to be considerably weak in hydrolyzing membrane phospholipids, although its activities were distinct. BP-I and BP-Ii share a common sequence with the sole exception of Asp-67 (BP-I) and Asn-67 (BP-II) in the aligned sequences. This implies that the amino acid at position 67 of Lys-49-PLA2s is the residue required for discriminatory recognition of β-arachidonoyl-phospholipid membranes.

AB - Basic proteins I and II (BP-I and BP-II) isolated from the venom of Trimeresurus flavoviridis (Habu snake) are isozymes of highly active Asp-49-phospholipase A2 (Asp-49-PLA2) and classified into the group Lys-49-PLA2. BP-II was found to elicit a strong contraction of Guinea pig ileum, and this activity was inhibited completely by 1 μM indomethacin, an inhibitor of the arachidonate cascade. BP-II was inactive in the Ca2+ -free medium, and p-bromophenacy lated His-48-BP-II was also inactive. BP-II exhibited no binding affinity for the cells expressinng PLA2 receptors. These results indicated that the contraction elicited by BP-II is due to the hydrolytic action of BP-II, liberating archidonic acid from the ileum phospholipid biomembranes. In spite of its limited lipolytic activities (av. 0.9% of Asp-49-PLA2) against monomers and micelles of synthetic phospholipids, BP-II hydrolyzed considerably strongly the phospholipids in the artificial bilayer vesicles. Arachidonic acid released from liposomes of β-arachidonoyl-γ-stearoyl-l-α-phosphatidylcholic was determined by HPLC, and the activity of BP-II was estimated to be about 75% as compared to Asp--49-PLA2. Liposomes encapsulating carboxyflurescein exhibited a strong dye-leakage induced by BP-II in a concentration-dependent manner, only in the Ca2+-containing buffer. The net result from all these observations was that BP-II, a Lys-49-PLA2, is an enzyme that hyrolyzes the membrane phospholipids. In contrast to BP-II, BP-I was found to be considerably weak in hydrolyzing membrane phospholipids, although its activities were distinct. BP-I and BP-Ii share a common sequence with the sole exception of Asp-67 (BP-I) and Asn-67 (BP-II) in the aligned sequences. This implies that the amino acid at position 67 of Lys-49-PLA2s is the residue required for discriminatory recognition of β-arachidonoyl-phospholipid membranes.

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