Studies on Biochemical Functions of Pyrroloquinoline Quinone

根据其定位和特性，将藜蛋白脱氢酶分为四类：(1)甲基营养菌体内的甲醇脱氢酶；(2)非甲基营养菌以乙醇为唯一碳源生长时的藜蛋白乙醇脱氢酶。(3)睾酮单胞菌以载脂蛋白形式形成醌血红蛋白醇脱氢酶，而恶臭假单胞菌以伯醇为碳源生长时形成holo酶。(4)形成醌血红蛋白醇脱氢酶-细胞色素c复合物，形成于乙酸细菌的细胞质膜内。在这一研究计划的过程中，以下新发现已被添加到明确的功能吡咯喹啉醌和醌蛋白。(1)膜结合、吡咯喹啉类依赖、乙醇脱氢酶在乙酸细菌呼吸链中起初级脱氢酶的作用。研究了从乙酰杆菌和亚羟葡萄糖杆菌中纯化的乙醇脱氢酶与泛醌直接反应的能力。乙醇氧化酶的重构是在室温下将无洗涤剂脱氢酶与泛醇氧化酶和含泛醌的磷脂通过辛基糖苷洗涤透析预先制备的蛋白脂质体结合。重组的蛋白脂质体具有合理水平的乙醇氧化酶活性，其电子转移反应也能产生膜电位。(2)研究了甲基营养乙酸杆菌（Acetobacter methanolicus）的甲醇和乙醇氧化酶呼吸链。在本研究中，在另一种类型的甲醇脱氢酶中发现了一个32 KDa的肽，该肽含有三个不同的亚基，其中两个对应于α亚基和β亚基。第3个亚基与反硝化副球菌（Paracoccus反硝化副球菌）或外链甲基杆菌（Methylobacterium extorquens AMl）的MoxJ基因同源性较高，最终确定为甲醇氧化酶产物。(3)从土壤中分离并鉴定的恶臭假单胞菌HK5可产生3种不同的藜蛋白醇脱氢酶作为全酶。当在乙醇上生长时，产生一种醌血红蛋白醇脱氢酶，而在甘油上生长时，产生另一种醌血红蛋白醇脱氢酶，可以氧化甘油。第三种类型的藜蛋白醇脱氢酶与在乙醇上生长的非甲基营养细菌中发现的相似。第三种醇脱氢酶不含血红素成分，不能以铁氰化钾作为电子受体氧化醇。少

The quinoprotein dehydrogenase has been classified into four groups according to their localization and characterization :(1) methanol dehydrogenase in methylotrophs,(2) quinoprotein alchold dehydrogenase from non-methylotrophic bacteria, when they are grown on ethanol as sole carbon source.(3) Quinohemoprotein alcohol dehydrogenase is formed as apo-enzyme in Commamonas testosteroni, while Pseudomonas putida forms holo-enzyme when they are grown on primary alcohols as the carbon source.(4) Quinohemoprotein alcohol dehydrogenase-cytochrome c complex is formed is formed in the cytoplasmic membranes of acetic acid bacteria.During the course of this research program, the following new findings have been added to clear up the function of pyrroloquinoline quinone and quinoproteins.(1) Membrane-bound, pyrroloquinoline quinone-dependent, alcohol dehydrogenase functions as the primary dehydrogenase in the respiratory chain of acetic acid bacteria. An ability of the enzyme to directly react with … More ubiquinone was investigated in alcohol dehydrogenases purified from both Acetobacter aceti and Gluconobacter suboxydans. The reconstitution of alcohol oxidase was done by binding the detergent-free dehydrogenase at room temperature to proteoliposomes that had been prepared in advance from a ubiquinol oxidase and phospholipids containing ubiquinone by detergent dialysis using octyl-glucoside. The proteoliposomes thus reconstituted had a reasonable level of ethanol oxidase activity, the electron transfer reaction of which was also able to generate a membrane potential.(2) Methanol and ethanol oxidase respiratory chains of the methylotrophic acetic acid bacterium, Acetobacter methanolicus, was investigated. In this study, an 32 KDa peptide was found in alternative type of methanol dehydrogenase which contained three different subunits two of which corresponded to alpha-subunit and beta-subunit. The third subunit was finally concluded to be the MoxJ product in methanol oxidase, because the N-terminal amino acid sequence of the peptide showed a high homology to that of MoxJ gene from Paracoccus denitrificans or Methylobacterium extorquens AMl.(3) Pseudomonas putida HK5, isolated from soil and identified thereafter, produced three different quinoprotein alcohol dehydrogenase as holo-enzyme. When grown on ethanol, it produced a quinohemoprotein alcohol dehydrogenase, While grown on glycerol, another quinohemoprotein alcohol dehydrogenase which can oxidize glycerol. The third type of quinoprotein alcohol dehyderogenase was similar to those found in non-methylotrophic bacteria which were grown on ethanol. The third alcohol dehydrogenase contained no heme component and was not able to oxidize alcohol with potassium ferricyanide as an electron acceptor. Less