Looking for evidence that flavin containing oxidase involves pyrroloquinoline quinone

几种含有黄素的氧化酶，如黄嘌呤氧化酶、d -氨基酸氧化酶、葡萄糖氧化酶和其他被认为含有黄素的酶，例如：尿酸酶或抗坏血酸氧化酶，通过蛋白水解和/或酸水解水解。对水解产物进行分析，看是否有任何信号显示吡咯喹啉醌（PQQ）或PQQ-发色团的存在。用Sephadex G-10色谱柱分离后，检测各部分作为载葡萄糖脱氢酶（EC 1.1.99.17）假体基的活性。从一些不涉及PQQ的参比蛋白，如溶菌酶和几种重结晶制剂的nadd依赖性醇脱氢酶的色谱图来看，PQQ没有活性。另一方面，尿酸酶、黄嘌呤氧化酶、抗坏血酸氧化酶的水解产物在其分子中明显显示PQQ的存在。这些水解液也可用作一些微生物的生长刺激物质，以减少其生长的滞后期。光谱数据表明PQQ或PQQ相关化合物可能存在于单个酶分子中。用PQQ和牛血清白蛋白进行交替模型试验。当PQQ与牛血清白蛋白在中性pH下混合时，观察到PQQ与血清白蛋白的自发结合。所得pq - bsa在色谱处理中相当稳定，在336 nm处有一个吸收峰。PQQ通过一个希夫碱与牛血清白蛋白赖氨酸残基的伊普西隆氨基结合。

Several flavin containing oxidases, such as xanthine oxidase, D-amino acid oxidase, glucose oxidase, and other enzymes which have been assumed to contain flavin, eg. uricase or ascorbate oxidase, were hydrolyzed by proteolysis and/or acid hydrolysis. The resulting hydrolysates were andlyzed to see whether exist any signal showing the presence of pyrroloquinoline quinone (PQQ) or PQQ-chromophores. After fractionation with Sephadex G-10 column, each fraction was examined of its activity as the prosthetic group for apo-glucose dehydrogenase (EC 1.1.99.17). From the chromatogram of some reference proteins in which PQQ is never involved, such as lysozyme and NAD-dependent alcohol dehydrogenase of several recrystallized preparations, showed no PQQ activity. On the other hand, of the examine candidates, the hydrolysate from uricase, xanthine oxidase, ascorbate oxidase potently showed the presence of PQQ in their molecule. These hydrolysate were also available as the growth stimulating substance for some microorganisms to reduce the lag phase in their growth. Spectroscopic data indicated the possibility of the existence of PQQ or PQQ-related compound in individual enzyme molecule.Alternative model experiment was carried out with PQQ and bovine serum albumin. When PQQ was mixed with bovine serum albumin at neutral pH, a spontaneous binding of PQQ to serum albumin was observed. The resulting PQQ-BSA was fairly stable for chromatographic treatments and exhibited absorption spectrum having a peak at 336 nm. PQQ was convinced to be asociated via ipsilon amino group of lysyl residue of BSA via a Schiff base.