Energy-dispersive x-Ray Analysis of Phosphorus, Potassium, Magnesium, and Calcium in Globoid Crystals in Protein Bodies from Different Regions of Cucurbita maxima Embryos

The seeds of Cucurbita maxima contain protein bodies with electrondense globoid crystals. Because of their density globoid crystals are ideal material for energy-dispersive x-ray (EDX) analysis studies of elemental composition. Fixation trials were carried out to test globoid crystal extraction during glutaraldehyde fixation, water washing, and ethanol dehydration. Glutaraldehyde fixation without subsequent washing or dehydration alone produced no significant changes in elemental composition of cotyledon globoid crystals. If glutaraldehyde fixation was followed by water washes or ethanol dehydration there was some loss of the major globoid crystal elements but the relative percentages of the elements P, K, Ca, and Mg remained relatively unchanged. In this paper results of a study of the P, K, Mg, and Ca content of globoid crystals in different tissues of squash embryos are presented. The globoid crystals in the radicle were found to be the least dense in the embryo. Globoid crystals from all embryo regions contained P, K, and Mg. In the various embryo regions P and Mg maintained relatively constant proportions of the globoid crystal composition while K and Ca varied. Of particular significance is the distribution of Ca which is generally an immobile element. Calcium was found in highest amounts in the globoid crystals of the radicle and stem regions while globoid crystals in much of the cotyledon contained little, if any, Ca. The Ca storage thus seems to be spatially arranged in a manner that would aid early growth of the root-shoot axis.     

油菜和玉米花粉粒中元素分布的研究

以油菜和玉米花粉为材料 ,应用扫描电镜及能谱仪技术对花粉中内外元素分布进行了初步研究。结果表明 :花粉粒萌发沟区与非萌发沟区 ,花粉壁与原生质内外的不同部位 ,Fe、Zn、Si、Al、Ca、P、S、Mg、K、Cl、Cu、Mn、Co及Ni1 4种元素组成有很大差异 ;两种花粉的萌发沟区都未检测出Fe元素 ,而非萌发沟区的Fe元素相对含量为 1 .1 2 %～ 3 68% ;花粉壁中比花粉原生质中更富含Si,Ca,K ,Cl等元素。本项研究为蜜源植物花粉的开发和利用提供了有价值的资料。     

Studies on Proteolysis in Stored Muscle

Changes in the nonprotein nitrogenous compounds produced from rabbit skeletal muscle (L. dorsi) by proteolysis were investigated.

The value of trichloroacetic acid soluble nitrogen, ninhydrin positive materials and phenol reagent positive materials increased during storage at low and high temperature. Changes in bound and free amino acid contents produced by proteolysis during storage were assayed by amino acid analyzer. Most of free amino acids except taurine increased remarkably. Amounts of asparatic acid, glutamic acid, glycine, β-alanine and histidine were increased after hydrolysis as compared with those before hydrolysis.

By using five kinds of Dowex 50 columns, changes in the distributive patterns of the nonprotein nitrogenous compounds were also studied.     

Studies on Proteolysis in Stored Muscle

Some physicochemical properties of the cathepsin D purified from the rabbit muscle (L. dorsi) were investigated.

The sedimentation coefficient (s_20,w) and the molecular weight determined from sedimentation equilibrium experiment was 3.83 S and 29,000~30,000, respectively.

The amino acid composition of the enzyme was determined with an automatic amino acid analyzer.

The proteolytic specificity of the enzyme was also investigated using the B-chain of oxidized beef insulin as the substrate. The cathepsin D cleaved the bonds Phe-Val, Ala-Leu, Leu-Tyr and Tyr-Leu. The specificity of the cathepsin D was fairly similar to that of the pepsin.     

Studies on Proteolysis in Stored Muscle

The amounts of proteolytic products derived from each fraction of rabbit muscle proteins by purified cathepsin D from rabbit muscle, and the effects of cathepsin D and pepsin treatments on ATPase activity of myosin B were studied.

Water-soluble protein was most rapidly hydrolyzed, followed by myosin A, actin, and myosin B.

In comparison with the control, the decrease of the Mg-enhanced ATPase activity of myosin B treated with cathepsin D was observed, but no difference was observed in the Ca-enhanced ATPase activity of myosin B by this treatment.

The removal of native tropomyosin in myosin B was not recognized by cathepsin D treatment, which was different from trypsin treatment.

The effect of pepsin treatment on the Mg-enhanced ATPase activity of myosin B was fairly similar to that of cathepsin D treatment.     

Dissection of Human Vitreous Body Elements for Proteomic Analysis

The vitreous is an optically clear, collagenous extracellular matrix that fills the inside of the eye and overlies the retina. ^1,2 Abnormal interactions between vitreous substructures and the retina underlie several vitreoretinal diseases, including retinal tear and detachment, macular pucker, macular hole, age-related macular degeneration, vitreomacular traction, proliferative vitreoretinopathy, proliferative diabetic retinopathy, and inherited vitreoretinopathies. ^1,2 The molecular composition of the vitreous substructures is not known. Since the vitreous body is transparent with limited surgical access, it has been difficult to study its substructures at the molecular level. We developed a method to separate and preserve these tissues for proteomic and biochemical analysis. The dissection technique in this experimental video shows how to isolate vitreous base, anterior hyaloid, vitreous core, and vitreous cortex from postmortem human eyes. One-dimensional SDS-PAGE analyses of each vitreous component showed that our dissection technique resulted in four unique protein profiles corresponding to each substructure of the human vitreous body. Identification of differentially compartmentalized proteins will reveal candidate molecules underlying various vitreoretinal diseases.     

Acid Carboxypeptidases in Grains and Leaves of Wheat, Triticum aestivum L

Extracts of resting and germinating (3 days at 20°C) wheat (Triticum aestivum L. cv Ruso) grains rapidly hydrolyzed various benzyloxycarbonyldipeptides (Z-dipeptides) at pH 4 to 6. Similar activities were present in extracts of mature flag leaves. Fractionation by chromatography on CM-cellulose and on Sephadex G-200 showed that the activities in germinating grains were due to five acid carboxypeptidases with different and complementary substrate specificities. The wheat enzymes appeared to correspond to the five acid carboxypeptidases present in germinating barley (L Mikola 1983 Biochim Biophys Acta 747: 241-252). The enzymes were designated wheat carboxypeptidases I to V and their best or most characteristic substrates and approximate molecular weights were: I, Z-Phe-Ala, 120,000; II, Z-Ala-Arg, 120,000; III, Z-Ala-Phe, 40,000; IV, Z-Pro-Ala, 165,000; and V, Z-Pro-Ala, 150,000. Resting grains contained carboxypeptidase II as a series of three isoenzymes and low activities of carboxypeptidases IV and V. During germination the activity of carboxypeptidase II decreased, those of carboxypeptidases IV and V increased, and high activities of carboxypeptidases I and III appeared. The flag leaves contained high activity of carboxypeptidase I and lower activities of carboxypeptidases II, IV, and V, whereas carboxypeptidase III was absent.     

Studies on Changes in Stored Shell Eggs

The intrinsic viscosity of ovomucin(B) from the thick white was higher than that of the thin white. The relative area of the fast moving component in the electrophoretic pattern of ovomucin(B) from the thick white was about twice that of the thin white. The hexose, hexosamine and sialic acid contents of ovomucin(B) from the thick white were higher than those of the thin white. The intrinsic viscosity and carbohydrate content of ovomucin from the egg white previously freed from lysozyme were low in comparison with those of ovomucin(B). The carbohydrate contents of crude lysozyme from the thick white were higher than those of the thin white, and crude lysozyme obtained from the thick white contained larger amounts of ovomucin-iike material than the thin white. From these results, discussion were made about the relation between the properties of ovomucin(B) and ovomucin-lysozyme interaction.     

Studies on Changes in Stored Shell Eggs

The content of the ovomucin gel obtained from the gel parts of stored thick white decreased during storage. Changes of the content of the ovomucin gel (A) was much larger than that of the ovomucin gel (B). The content of the ovomucin sol obtained from the sol parts of stored thick white increased during storage.

The hexose and hexosamine contents of the ovomucin gel (B) decreased to about one half and the sialic acid content decreased to one eighth after 20 days storage at 30°C On the other hand, the carbonhydrate contents of the ovomucin sol (B) increased during storage and those obtained from sol parts of the stored (20 days) thick white were higher than those of the control ovomucin gel (B) obtained from the newly laid thick white. The amino acid composition of the ovomucin gel (B) and sol (B) did not show a great deal of change during storage.

It is suggested from these results that the properties of the ovomucin gel (B) changed greatly during storage; one portion of the ovomucin gel (B), the carbohydrate-rich component, solubilized to the sol parts of stored thick white and the other portion, the carbohydrate-poor component, remained insoluble.     

Studies on Changes in Stored Shell Eggs

Being solubilized by treatment with 0.01 m mercaptoethanol, the ovomucin gel(B) was found in free boundary electrophoresis to contain subunits which were consisted of two components. Changes in the physicochemical properties of all the insoluble ovomucin gel(B) and sol(B) obtained from stored egg white were studied after this treatment.

The fast moving component of the ovomucin gel(B) in free boundary electrophoresis decreased during storage and disappeared completely after 30 days. On the other hand, the fast moving component of the ovomucin sol(B) increased during storage.

The acid mucoprotein concentration of the ovomucin gel(B) in acrylamide gel electrophoresis decreased and that of the ovomucin sol(B) increased during storage, although the protein pattern did not show significant changes.

The interaction of the ovomucin gel(B) with lysozyme decreased whereas that of the ovomucin sol(B) increased during storage.

By summarizing these results, a model of ovomucin gel structure and a mechanism of egg white thinning were proposed.     

水稻强弱势籽粒核酸和蛋白质含量的差异

水稻抽穗当天,强势粒(上部一次枝梗籽粒)子房中总RNA、mRNA含量都高于弱势粒(下部二次枝梗籽粒),蛋白质含量却低于弱势粒.抽穗后第5天,强势粒子房中总RNA含量仍然高于弱势粒,mRNA含量两者差异不大,蛋白质含量依然低于弱势粒.抽穗后0～5d,强势粒子房中总RNA、mRNA含量变化不大,蛋白质含量下降显著;弱势粒子房中总RNA含量上升显著,mRNA含量、蛋白质含量变化不大.抽穗当天和抽穗后第5天,强势粒子房中有一些小分子rRNA带出现;弱势粒则相反,小分子rRNA基本上看不见.     

小麦籽粒内AGPase质体型小亚基的克隆和序列分析

文章从小麦籽粒中克隆了淀粉合成关键酶AGPase的质体型小亚基(SSU II)的cDNA序列.其中间部分与3'端序列与大麦SSU II(Z48563)的同源性高达97%,但在5'端特异的转移肽切割位点却比Z48563缺失一段113 bp序列.本文克隆的SSU II其特异5'端序列与已报道的大麦(Z48563)、小麦(536819)、玉米(AF334960)进行多序列比对和同源性比较显示,它们的亲缘关系较远,表明这可能是一个新的SSU II基因.另外,在检测的22个现今推广面积较大的小麦品种中,SSU II 5'端缺失序列的现象都普遍存在.     

Ultrastructure of Endosperm Protein Bodies in Developing Rice Grains Differing in Protein Content

Ultrastructural aspects of the development of protein bodies(aleurone grains) in endosperm of grains of two rices differingin protein content are described. Formation of rough endoplasmicreticulum complexes prior to protein deposition was observedonly in the higher protein grain. From 8 days after floweringthree types of protein body were observed, one of which wasrestricted to the peripheral endosperm (sub-aleurone) layers.The higher protein grain had a greater number of protein bodiesand rough endoplasmic reticulum in the endosperm cells thanthe lower protein grain. Increase in total protein with maturitywas the result of increased number of protein bodies ratherthan increase in size; the protein bodies were concentratedin the peripheral endosperm layers.     

Changes in Abscisic Acid Levels in Developing Grains of Wheat (Triticum aestivum L.

ABA has been determined in wheat grains during their development. The maximum level of ABA occurred approximately 7 weeks afterear emergence (40 d after peak anthesis). This level subsequentlyfell sharply as a result of metabolism of the ABA by the maturinggrains.     

Spectroscopic Imaging of Protein Crystals in Crystallization Drops

Automatic imaging and scoring of crystallization drops is an essential step in high-throughput crystallography. Presently, white-light images of crystallization drops are acquired robotically and the images are analyzed and scored using pattern recognition algorithms. However, the scoring part remains unreliable as crystals and microcrystals are not always recognized by existing feature-extraction and recognition algorithms. We propose a fundamental shift in crystal monitoring through spectroscopic imaging of crystallization drops. This method converts the problem of automatic crystal detection from one of pattern recognition into one of intensity (concentration) analysis. The latter can be more robust and reliable.     

Histochemical Localization of Calcium Oxalate Crystals in Starch Grains of Yams (Dioscorea)

The bulbils and/or tubers of seven species of yams (Dioscorea)were examined for crystal content using light microscopy andhistochemistry. Calcium oxalate crystals in the form of raphide bundles werelocalized in the parenchymatous tissues. Within starch grains,crystals of various shapes and sizes were observed. The variationin shape and sizes of the intra-amylar crystals could be exploitedfor taxonomic purposes. Calcium oxalate crystals appear to serve a storage functionin these starch grains. Yams, Dioscorea, raphides, oxalate crystals, histochemistry     

Protein Electrophoresis of Single Pollen Grains of Hibiscus rosa-sinensis

Electrophoresis and staining of proteins from the single pollen grains of Hibiscus rosasinensis have been developed by using general ultrathin polyacrylamide gel combined with highly-sensitive silver staining technique. The result revealed that the pollen abortion could occur in different stages of pollen development. The protein patterns varied greatly in different stages of pollen development, even in the different pollen grains in the same anther at the same development stage. Some bands exhibited a disjunction by classical Mendelian ratio 1: 1, suggesting that the gene loci were heterogeneous and the proteins were related to the expression of the genes at the early stage of pollen development.     

Ultrastructure of Protein Crystals in Potato and Tomato Trichomes

Large protein crystals were located in the leaf and stem trichomesof Solanum tuberosum L. and Lycopersicon esculentum Mill. Inpotato the crystals ranged from 1.05 to 4.5 µm (average2.3 µm) on a side and in tomato they ranged from 1.16to 3.5 µm (average 2.7 µm) on a side. The proteinnature of the crystals was determined by histochemical stainingwith Coumassie brilliant blue R250 and aniline blue black. Thecrystalline structure of the inclusions was observed in ultrathinsections using electron microscopy. In potato, in cleared areasof the cytoplasm, ribosomes were observed scattered among proteinfilaments. The filaments were approximately 7 nm in diameter.Morphologically similar crystals were observed in the tomatotrichomes but the protein filaments were smaller (approximately4 nm in diameter). Protein crystals were also observed in palisadeand spongy parenchyma and epidermal leaf cells in tomato. Protein crystals, trichomes, potato, Solanum tuberosum L., tomato, Lycopersicon esculentum Mill., ultrastructure     

固氮酶锰铁蛋白的晶体生长

从以Mn代钼的固氮培养基中固氮生长的固氮菌（Azotobacter vinelandii Lipmann）突变种UW3中分离纯化的MnFe蛋白,在一定的结晶条件下,可从溶液中析出深棕色的短斜四棱柱晶体。Tris和Hepes缓冲液、NaCl、MgCl2和PEG的浓度及结晶方法等,对该蛋白的出晶率、晶核数目、晶体大小和质量均有明显的影响,PEG浓度的改变还可使该蛋白晶体的晶型发生变化。MnFe蛋白结晶所需的上述化合物的最适浓度与缺失nifZ固氮菌突变种△nifZ MoFe需的最适浓度有所不同。ＳＤＳ凝胶电泳表明,晶体溶解的蛋白与结晶前的MnFe蛋白基本相同。结果表明：该晶体为MnFe蛋白的晶体。