Chemical cross-linking study of complex formation between methylamine dehydrogenase and amicyanin from Paracoccus denitrificans.
Chemical cross-linking study of complex formation between methylamine dehydrogenase and amicyanin from Paracoccus denitrificans.
复制标题
脱氮副球菌甲胺脱氢酶与阿霉素之间形成复合物的化学交联研究。
DOI：
10.1021/bi00474a012
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发表时间：
1990
期刊：
影响因子：
2.9
通讯作者：
Davidson,VL
中科院分区：
文献类型：
--
作者：
Kumar,MA;Davidson,VL
密西西比大学医学中心生物化学系，2500 N。州街，杰克逊，密西西比39216-4505，1989年11月20日接收; 1990年1月24日接收的修订的Mandalpt摘要：来自副球菌的两种可溶性周质氧化还原蛋白，醌蛋白甲胺脱氢酶和铜蛋白amicyanin，形成弱缔合的复合物，该复合物对于它们在电子传递中的生理功能是关键的[Gray，KA，Davidson，V.L.，& Knaff，D. B。（1988）J.Biol.Chem.263，13987-13990]，已经通过使用水溶性交联剂1-乙基-3-[3-（二甲基-氨基）丙基]碳二亚胺（EDC）研究了甲胺氧化酶和amicyanin之间的特异性相互作用。单独用EDC处理甲胺脱氢酶不引起分子间交联，但确实引起该α 2 β 2寡聚酶的分子内交联。形成的初级产物含有一个大亚基和一个小亚基。甲胺脱氢酶和amicyanin共价交联在EDC的存在下，形成至少两个不同的物种，这是通过非变性聚丙烯酰胺凝胶电泳（PAGE）进行鉴定。这些交联物质的形成依赖于离子强度，并且离子强度依赖性在pH 6.5下比在pH 7.5下大得多。不同的交联产物，pH值和离子强度的影响是不同的。这些交联的物种的SDS-PAGE和Western印迹分析表明，为amicyanin的相互作用的主要网站是大亚基的甲胺脱氢酶，这种协会可以通过疏水相互作用稳定。根据这些结果，提出了一个方案的amicyanin与甲胺脱氢酶的相互作用，这是与以前的数据一致的物理，动力学和氧化还原特性的这种复杂的。
Department of Biochemistry, The University of Mississippi Medical Center, 2500 N. State St., Jackson, Mississippi 39216-4505 Received November 20, 1989; Revised Manuscript Received January 24, 1990 abstract: Two soluble periplasmic redox proteins from Paracoccus denitrificans, the quinoprotein me-thylamine dehydrogenase and the copper protein amicyanin, form a weakly associated complex that is critical to their physiological function inelectron transport [Gray, KA, Davidson, V. L., & Knaff, D. B.(1988) J. Biol. Chem. 263, 13987-13990], The specific interactions between methylamine dehydrogenaseand amicyanin have been studied by using the water-soluble cross-linking agent 1-ethyl-3-[3-(dimethyl-amino) propyl] carbodiimide (EDC). Treatment of methylamine dehydrogenase alone with EDC caused no intermolecular cross-linking but did cause intramolecular cross-linking of this a2 (32 oligomeric enzyme. The primary product that was formed contained one large and one small subunit. Methylamine dehydrogenase and amicyanin were covalently cross-linked in the presence of EDC to form at least two distinct species, which were identified by nondenaturing polyacrylamide gel electrophoresis (PAGE). The formation of these cross-linked species was dependent on ionic strength, and the ionic strength dependence was much greater at pH 6.5 than at pH 7.5. The effects of pH and ionic strength were different for the different cross-linked products. SDS-PAGE and Western blot analysis of these cross-linked species indicated that the primary site of interaction for amicyanin was the large subunit of methylamine dehydrogenase and that this association could be stabilized by hydrophobic interactions. In light of these results a scheme is proposed for the interaction of amicyanin with methylamine dehydrogenase that is consistent with previous data on the physical, kinetic, and redox properties of this complex.