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広島大学水畜産学部紀要 Volume 12 Issue 2
published_at 1973-12-20
紅藻オキツノリ抽出液による塩基性アミノ酸の酵素的分解
Enzymatic Decomposition of Basic Amino Acids by the Cell Free Extract of a Red Alga， Gymnogongrus flabelliformis
Ito Keijl
Miyazawa Keisuke
Abstract
紅藻の一種オキツノリGymnogongrus flabelliformisを-200℃に凍結貯蔵すると藻体遊離アミノ酸の主要成分であるgigartinineおよびcitrullineが著しく減少し7週間の貯蔵で両者とも95%以上が分解された.藻体をpH8.0のTris緩衝液とともに磨砕して得た抽出液を25℃に置いた場合には更に顕著で、あって，gigartinineは3時間で完全に消失した.更に透析処理した抽出液に17種のアミノ酸混合液を添加して分解性をみたところ，この抽出液は特に塩基性アミノ酸を強く分解することを知った.
以上のことは本海藻に塩基性アミノ酸を特異的に分解する酵素の存在を示唆したので，この反応に対する緩衝液の種類の影響，至適温度，pH，基質特異性，分解生産物について調べた.結果は以下の如くである.
(1) 燐酸緩衝液はTris緩衝液.硼砂緩衝液に比較してarginine分解の活性を低下させる.
(2) 反応の至適温度は30℃附近にある.
(3) 反応の至適pHは8.0～9.0の間にある.
(4) 塩基性アミノ酸であるL-arginine ，L-ornithine， L-lysine，L-gigartinineは強く分解するが，L-histidine L-methionineにも弱い分解力を持つ.DL-Ornithine，DL-lysine，DL-gigartinineは50%しか分解されず，この反応はL-アミノ酸に特異的であると推察される.
(5) Arginine，gigartinineの分解生産物をペーパークロマトグラフィーによって検索したところ，いずれも主生成物として相当するα-ケト酸が検出された.したがって，この海藻抽出液には塩基性アミノ酸の酸化的脱アミノ反応に関与する酵素の存在が推定された.
以上のことは本海藻に塩基性アミノ酸を特異的に分解する酵素の存在を示唆したので，この反応に対する緩衝液の種類の影響，至適温度，pH，基質特異性，分解生産物について調べた.結果は以下の如くである.
(1) 燐酸緩衝液はTris緩衝液.硼砂緩衝液に比較してarginine分解の活性を低下させる.
(2) 反応の至適温度は30℃附近にある.
(3) 反応の至適pHは8.0～9.0の間にある.
(4) 塩基性アミノ酸であるL-arginine ，L-ornithine， L-lysine，L-gigartinineは強く分解するが，L-histidine L-methionineにも弱い分解力を持つ.DL-Ornithine，DL-lysine，DL-gigartinineは50%しか分解されず，この反応はL-アミノ酸に特異的であると推察される.
(5) Arginine，gigartinineの分解生産物をペーパークロマトグラフィーによって検索したところ，いずれも主生成物として相当するα-ケト酸が検出された.したがって，この海藻抽出液には塩基性アミノ酸の酸化的脱アミノ反応に関与する酵素の存在が推定された.
Abstract
The red alga, Gymnogongrus flabelliformis has a peculiar free amino acid pattern, in which one characteristic amino acid, gigartinine, is the most dominant component1-4). Recently, we observed that the contents of gigartinine and citrulline in the extracts of the alga decreased markedly during storage at about -20℃.
The purpose of this paper is to report that the cell free extract of Gymnogongrus flabelliformis has an enzymatic activity for the decomposition of basic L-amino acids.
1) The cell free extract was prepared by grinding the alga with an equal volume of pH 8.0 Tris buffer in a mortar previously chilled and centrifuging for 30 min. at 10,000 rpm under chilling. The supernatant obtained was dialyzed against distilled water for 24 hours.
2) The extract showed a deaminative activity towards L-gigartinine, L-arginine, L-lysine and L-ornithine, and decomposed 50% of DL-gigartinine, DL-lysine and DL-ornithine. It suggests that D-isomers are not affected.
3) The influence of the different kinds of buffer solution on the decomposing activity of the extract towards arginine was examined. The phosphate buffer showed a slightly inhibitive effect.
4) Effects of pH and temperature on the decomposing activity of the extract were examined. The pH optimum was relatively broad. The activity of the extract did not vary appreciably from pH 8.0 to 9.0. The optimum temperature was about 30℃.
5) The decomposition products of arginine and gigartinine were surveyed by paperchromatography and the corresponding a-keto acid to each one of the amino acids was detected, respectively. The data suggest that the extract of the alga has an activity to catalyze oxidative deamination of basic L-amino acids.
The purpose of this paper is to report that the cell free extract of Gymnogongrus flabelliformis has an enzymatic activity for the decomposition of basic L-amino acids.
1) The cell free extract was prepared by grinding the alga with an equal volume of pH 8.0 Tris buffer in a mortar previously chilled and centrifuging for 30 min. at 10,000 rpm under chilling. The supernatant obtained was dialyzed against distilled water for 24 hours.
2) The extract showed a deaminative activity towards L-gigartinine, L-arginine, L-lysine and L-ornithine, and decomposed 50% of DL-gigartinine, DL-lysine and DL-ornithine. It suggests that D-isomers are not affected.
3) The influence of the different kinds of buffer solution on the decomposing activity of the extract towards arginine was examined. The phosphate buffer showed a slightly inhibitive effect.
4) Effects of pH and temperature on the decomposing activity of the extract were examined. The pH optimum was relatively broad. The activity of the extract did not vary appreciably from pH 8.0 to 9.0. The optimum temperature was about 30℃.
5) The decomposition products of arginine and gigartinine were surveyed by paperchromatography and the corresponding a-keto acid to each one of the amino acids was detected, respectively. The data suggest that the extract of the alga has an activity to catalyze oxidative deamination of basic L-amino acids.
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