Haldar U C, Saha S K, Beavis R C, Sinha N K
Department of Chemistry, Bose Institute, Calcutta, India.
J Protein Chem. 1996 Feb;15(2):177-84. doi: 10.1007/BF01887398.
Two trypsin inhibitors, LA-1 and LA-2, have been isolated from ridged gourd (Luffa acutangula Linn.) seeds and purified to homogeneity by gel filtration followed by ion-exchange chromatography. The isoelectric point is at pH 4.55 for LA-1 and at pH 5.85 for LA-2. The Stokes radius of each inhibitor is 11.4 A. The fluorescence emission spectrum of each inhibitor is similar to that of the free tyrosine. The biomolecular rate constant of acrylamide quenching is 1.0 x 10(9) M-1 sec-1 for LA-1 and 0.8 x 10(9) M-1 sec-1 for LA-2 and that of K2HPO4 quenching is 1.6 x 10(11) M-1 sec-1 for LA-1 and 1.2 x 10(11) M-1 sec-1 for LA-2. Analysis of the circular dichroic spectra yields 40% alpha-helix and 60% beta-turn for La-1 and 45% alpha-helix and 55% beta-turn for LA-2. Inhibitors LA-1 and LA-2 consist of 28 and 29 amino acid residues, respectively. They lack threonine, alanine, valine, and tryptophan. Both inhibitors strongly inhibit trypsin by forming enzyme-inhibitor complexes at a molar ratio of unity. A chemical modification study suggests the involvement of arginine of LA-1 and lysine of LA-2 in their reactive sites. The inhibitors are very similar in their amino acid sequences, and show sequence homology with other squash family inhibitors.
已从棱角丝瓜（Luffa acutangula Linn.）种子中分离出两种胰蛋白酶抑制剂LA - 1和LA - 2，并通过凝胶过滤继以离子交换色谱法将其纯化至同质。LA - 1的等电点为pH 4.55，LA - 2的等电点为pH 5.85。每种抑制剂的斯托克斯半径为11.4 Å。每种抑制剂的荧光发射光谱与游离酪氨酸的相似。对于LA - 1，丙烯酰胺猝灭的双分子速率常数为1.0×10⁹ M⁻¹ s⁻¹，对于LA - 2为0.8×10⁹ M⁻¹ s⁻¹；对于LA - 1，K₂HPO₄猝灭的双分子速率常数为1.6×10¹¹ M⁻¹ s⁻¹，对于LA - 2为1.2×10¹¹ M⁻¹ s⁻¹。圆二色光谱分析得出，LA - 1含有40%的α - 螺旋和60%的β - 转角，LA - 2含有45%的α - 螺旋和55%的β - 转角。抑制剂LA - 1和LA - 2分别由28和29个氨基酸残基组成。它们不含苏氨酸、丙氨酸、缬氨酸和色氨酸。两种抑制剂均通过以1:1的摩尔比形成酶 - 抑制剂复合物来强烈抑制胰蛋白酶。化学修饰研究表明，LA - 1的精氨酸和LA - 2的赖氨酸参与了它们的活性位点。这两种抑制剂的氨基酸序列非常相似，并与其他南瓜科抑制剂显示出序列同源性。
