Periodate oxidation of sperm-whale myoglobin and the role of the methionine residues in the antigen-antibody reaction.
Periodate oxidation of sperm-whale myoglobin and the role of the methionine residues in the antigen-antibody reaction.
复制标题
抹香鲸肌红蛋白的高碘酸盐氧化以及蛋氨酸残基在抗原抗体反应中的作用。
作者：
M. Atassi
1.研究了高碘酸盐对抹香鲸高铁肌红蛋白及其载脂蛋白的氧化反应，在不同的pH和温度条件下，寻找试剂具有特异性的条件。2.在pH6.8和22 ℃时，高碘酸盐的消耗在每摩尔肌红蛋白43摩尔高碘酸盐时在3（1/2）小时内停止。两个甲硫氨酸残基，两个色氨酸残基，三个酪氨酸残基和两个组氨酸残基被氧化;丝氨酸在水解产物中从6个增加到9个残基/mol。3.在pH5.0和22 ℃下，消耗在4（1/2）小时内趋于稳定，每摩尔肌红蛋白消耗26摩尔高碘酸盐，导致两个甲硫氨酸残基、两个色氨酸残基、三个酪氨酸残基和两个组氨酸残基发生修饰;丝氨酸从6个残基/摩尔增加到7个残基/摩尔。并且铁血红素也遭受了相当大的氧化。4.在pH5.0和0度下氧化完成（4小时）。在22摩尔高碘酸盐/摩尔肌红蛋白的消耗和在甲硫氨酸、酪氨酸和色氨酸残基的修饰中。光谱研究表明血红素基团氧化。该衍生物与兔抗MbX（通过CM-纤维素色谱法获得的主要组分编号10; Atassi，1964）的抗血清反应非常差。5.在pH5.0和0 ℃条件下，脱辅基肌红蛋白的氧化在4 h内完成。消耗7.23摩尔高碘酸盐/摩尔脱辅基蛋白。氧化速率按降序排列为：蛋氨酸;色氨酸;酪氨酸; 7小时后。的反应的以下残留物/mol。氧化：蛋氨酸，2.0;色氨酸，1.6;酪氨酸，0.99。没有肽键被切割。从7小时制备的高铁肌红蛋白。氧化的载脂蛋白显示与MbX的抗血清的反应性已显著降低。6.脱辅基蛋白的温和氧化（2摩尔过量的高碘酸盐，pH5.0，0度，2小时）结果表明，在1.66个蛋氨酸残基/mol的条件下，从该脱辅基蛋白制备的高铁肌红蛋白在光谱、电泳和免疫化学上与天然MbX相同。结果表明，55和131位的蛋氨酸残基不是高铁肌红蛋白抗原位点的必需部分。
1. Oxidation of sperm-whale metmyoglobin and its apoprotein with periodate has been investigated under various conditions of pH and temperature to find those under which the reagent acted with specificity. 2. At pH6.8 and 22 degrees consumption of periodate ceased in 3(1/2)hr. at 43 moles of periodate/mole of myoglobin. The two methionine residues, the two tryptophan residues, the three tyrosine residues and two histidine residues were oxidized; serine increased in the hydrolysates from 6 to 9 residues/mol. 3. At pH5.0 and 22 degrees , consumption levelled off in 4(1/2)hr. at 26 moles of periodate/mole of myoglobin and resulted in the modification of the two methionine residues, the two tryptophan residues, the three tyrosine residues and two histidine residues; serine increased from 6 to 7 residues/mol. and, also, ferrihaem suffered considerable oxidation. 4. Oxidation at pH5.0 and 0 degrees resulted at completion (4hr.) in the consumption of 22 moles of periodate/mole of myoglobin and in the modification of the methionine, tyrosine and tryptophan residues. Spectral studies indicated oxidation of the haem group. This derivative reacted very poorly with rabbit antisera to MbX (the major component no. 10 obtained by CM-cellulose chromatography; Atassi, 1964). 5. Oxidation of apomyoglobin at pH5.0 and 0 degrees was complete in 4hr. with the consumption of 7.23 moles of periodate/mole of apoprotein. The rate of oxidation in decreasing order was: methionine; tryptophan; tyrosine; and after 7hr. of reaction the following residues/mol. were oxidized: methionine, 2.0; tryptophan, 1.6; tyrosine, 0.99. No peptide bonds were cleaved. Metmyoglobin prepared from the 7hr.-oxidized apoprotein showed that the reactivity with antisera to MbX had diminished considerably. 6. Milder oxidation of apoprotein (2 molar excess of periodate, pH5.0, 0 degrees , 2hr.) resulted in the modification of 1.66 residues of methionine/mol. Metmyoglobin prepared from this apoprotein was identical with native MbX spectrally, electrophoretically and immunochemically. It was concluded that the methionine residues at positions 55 and 131 were not essential parts of the antigenic sites of metmyoglobin.