Unfolding intermediate in the peroxisomal flavoprotein D-amino acid oxidase

The flavoenzyme d-amino acid oxidase (DAAO) from Rhodotorula gracilis is a peroxisomal enzyme and a prototypical member of the glutathione reductase family of flavoproteins. DAAO is a stable homodimer with a FAD molecule tightly bound to each 40-kDa subunit. In this work, the urea-induced unfolding of dimeric DAAO was compared with that of a monomeric form of the same protein, a deleted dimerization loop mutant. By using circular dichroism spectroscopy, protein and flavin fluorescence, 1,8-anilinonaphtalene sulfonic acid binding and activity assays, we demonstrated that the urea-induced unfolding of DAAO is a three-state process, yielding an intermediate, and that this process is reversible. The intermediate species lacks the catalytic activity and the characteristic tertiary structure of native DAAO but has significant secondary structure and retains flavin binding. Unfolding of DAAO proceeds through formation of an expanded, partially unfolded inactive intermediate, characterized by low solubility, by increased exposure of hydrophobic surfaces, and by increased sensitivity to trypsin of the beta-strand F5 belonging to the FAD binding domain. The oligomeric state does not modify the inferred folding process. The strand F5 is in contact with the C-terminal alpha-helix containing the Ser-Lys-Leu sequence corresponding to the type 1 peroxisomal targeting signal, and this structural element interacts with the N-terminal betaalphabeta flavin binding motif (Rossmann fold). The expanded conformation of the folding intermediate (and in particular the higher disorder of the mentioned secondary structure elements) could match the structure of the inactive holoenzyme required for in vivo trafficking of DAAO through the peroxisomal membrane.     

Why oxalic acid protects cellulose during ozone treatments?

This study shows that oxalic acid is a very effective additive for the ozone treatments compared with other additives. The required concentration is very small and even lower than the concentration formed during the stage itself. Several different mechanisms for the action of oxalic acid as an additive during the ozone bleaching are examined. He is advisable to have a pH acid during Z stage, but it has been proved that pH value is not the only factor that positively influences. Oxalic acid has an additional effect due to different factors that increase the selectivity of the Z stage. This acid has a minor ^oOH scavenging capacity than others acids, however, it prevents the cellulose degradation to a greater extent. Moreover, less ^oOH are formed when oxalic acid is added. Oxalic acid decreases swelling of cellulose and stabilises ozone avoiding its decomposition, then ozone is not consumed by oxalic acid. Metal ions, which are detrimental during ozone stage, can be chelating by oxalic acid. The most interesting outcome is that oxalic acid could act as hydrogen donor inhibiting the formation of ^oOH radicals. Thus, in studying the effects of oxalic acid, we have determined the requisites for an additive to be optimally effective to protect cellulose during ozone treatments.     

Production of oxalic acid by some fungi infected tubers

Oxalic acid (as oxalate) was detected in four tubers commonly used for food in Nigeria-Dioscorea rotundata (White yam), Solanum tuberosum (Irish potato), Ipomoea batatas (Sweet potato), and Manihot esculenta (cassava). Whereas healthy I. batata had the highest oxalic acid content, healthy M. esculenta contained the lowest. When all tubers were artifically inoculated with four fungi-Penicillium oxalicum CURIE and THOM, Aspergillus niger VAN TIEGH, A. flavus and A. tamarii KITA, there was an increase in oxalate content/g of tuber tissue. The greatest amount of oxalate was produced by P. oxalicum in D. rotundata tuber. Consistently higher amounts of oxalate were produced by the four fungi in infected sweet potato tuber than in any other tuber and consistently lower amounts of oxalate were produced by the four fungi in Irish potato tuber. Differences in the carbohydrate type present in the tubers and in the biosynthesis pathway are thought to be responsible for variation in the production of oxalate in the different tubers by the four fungi used.     

The photosynthetic production of oxalic acid in Oxalis corniculata

The biogenesis of oxalic acid in Oxalis corniculata has beeninvestigated. In O. corniculata the bulk of the oxalic acidis produced by CO₂ fixation both in light and in darkness butthe rate of its photosynthetic formation is much higher thanin darkness. Several other plants some of which are known toaccumulate oxalic acid e.g., Biophytum sensitivum, Averrhoacarambola, Impatiens balsamina, Amorphophallus campanulatusand Colocassia antiquorum also fix ¹⁴CO₂ into oxalic acid photosyntheticallywithin 1 min of exposure to the gas. In O. corniculata ¹⁴C canbe detected in oxalic acid within 5 sec and about half of thetotal ¹⁴C fixed in the 70% ethanol soluble fraction can be locatedin this compound after 5 min. This is accompanied by a declineof radioactivity in two compounds, the chromatographic behaviourand melting points of one of which and its DNP hydrazone aresimilar to those of an authentic sample of glyoxylic acid. Whenglyoxylate 1, 2-¹⁴C is incubated with Oxalis leaf homogenateit is converted to oxalate-¹⁴C. Glycolate is also metabolizedto oxalate. The conversion of both glycolate and glyoxylateare favoured by light. The C₂ compounds acetate and glycinehowever are utilized rather poorly. Sucrose-¹⁴C is also notmetabolized markedly for this purpose. (Received August 20, 1969; )     

Determination of plasma oxalic acid by high-pressure liquid chromatography

A new specific and sensitive method for determination of oxalic acid in plasma by High Performance Liquid Chromatography (HPLC) is described. The plasma sample is deproteinized by ultrafiltration. The oxalic acid in the ultrafiltrate is purified by precipitation with CaCl2, new dilution of calcium oxalate precipitate, oxalic acid extraction with diethyl-ether and total dryness of the sample. The losses of oxalic acid during this process are evaluated by the addition of oxalic acid (U-14C) before the precipitation step. The dried samples are redissolved in mobile phase (o-H3PO4, 0.05 M) and injected into a HPLC chromatograph, with reversed phase column (Lichrosorb RP-8, Merck). Oxalate peak is detected spectrophotometrically at 220 nm with a retention time of 3.20 minutes. The method shows a mean recovery value of 82.11, with an intra-run and between-run CV values of 2.54 and 6.95 respectively. The oxalic acid measured in plasma by this method is 291 +/- 89 micrograms/100 ml plasma ultrafiltrate, in 16 normal subjects.     

苔藓

闻名久远的五大连池．除了奇特的火山地貌．神奇的矿泉水之外还有着独特的高等与低等共存的植被．高大粗壮的树木统称乔木,较矮小丛生的木本植物被称为灌木．五彩缤纷的花草攀援的藤本但还有一些很不起眼的小型绿色苔藓植物这些苔藓植物多数形态矮小．一般高为数厘米至数十厘米直立丛生或匍匐生长多数种类属于喜湿性适应性强分布广是苔藓植物的特性。     

U-1分解草酸的检测及其降低草酸毒性的研究

草酸是油菜菌核病病菌(Sclerotinia sclerotiorum)致病的重要因子。以实验室分离得到的草酸分解菌U-1为材料,研究U-1分解草酸的能力及其降低草酸对油菜毒性的作用机理。检测U-1分解草酸的能力,即以草酸、U-1 草酸分别培养24 h后,利用高效液相色谱仪检测草酸的含量;研究U-1抑制草酸对油菜毒害的作用机理,即分别以清水、U-1、草酸、U-1 草酸处理油菜叶片,检测不同处理对油菜叶片中防卫反应0相关酶(PAL酶、PPO酶和PO酶)变化的影响。研究结果表明,与草酸单独处理组相比,U-1 草酸处理组草酸含量显著降低;U-1可以显著抑制草酸对油菜中PAL酶、PPO酶和PO酶活性的作用。     

植物病原真菌致病毒素草酸的研究进展

许多植物病原菌可以分泌草酸,草酸作为致病的关键因子在病原菌的侵染过程中发挥着重要作用,并与病原菌的致病性、毒性有密切关系。草酸可通过氧化和脱羧两条途径进行降解,因此可以将草酸降解酶基因导入植物,从而获得对这类病害的抗性。