Electronmicroscopic histochemical investigation of the surface of the cell membrane of the renal tubules with the use of ruthenium red and lantanum nitrate.

The renal tubules were investigated with the use of ruthenium red and lanthanum nitrate to titrate the carbohydrate groups which are localized on the differentiated surface of the cell membranes and intercellular spaces. Ruthenium red visualized the surface coat which is tightly bound to the outer lamellae of the cell membrane. Lanthanum nitrate used in this investigations is a valuable marker of the intercellular spaces. The applied markers have visualized in the kidney the canals which are originated from the foldings of membranes of the tubular cells which adhere to the basal membrane. The markers used in combination with the fixative glutaraldehyde-paraformaldehyde give an electrondense envelope of the cell membrane which is more distinct as compared with specimen treated with glutaraldehyde and markers only.     

The fine structure of campaniform sensilla on the halteres of Drosophila melanogaster

The campaniform sensilla on halteres of Drosophila were studied by electron microscopy in order to establish the relationships of functional elements in the sensory system. The surface of the sensillum consists of an oval cuticular cap membrane which may contain resilin, the rubberlike protein. A border of denser cuticle rings the cap membrane, and extending down around the neural process is a third type of cuticle filled with a fourth light fibrous type. The four cuticular components form a system for displacement of the neural process. The neural process is differentiated into a terminal fan-shaped structure projecting from a bulbous dilatation which tapers to a neck region ending proximally with two basal bodies. The neural process is packed with microtubules. Surrounding the dendrite is an inner enveloping cell, attached to the basal body region by septate desmosomes and by desmosomes to which microtubules of the enveloping cell are applied. An outer enveloping cell surrounds the inner one. The tip of the neural process is covered with a dense secretion which is tightly bound to the cap membrane. The dense secretion is surrounded by an extracellular fluid which might be compressed hydraulically by the cuticular system. The stimulus of cuticular distortion could thus be transmitted to the neural process which may be displaced between its fixed ends.     

Dependence of synergism of the combined effect of ultrasound and hyperthermia upon intensity of ultrasound

The inactivation of wild-type yeast Saccharomyces cerevisiae was studied after simultaneous treatment with ultrasound and hyperthermia. A temperature range was established within which ultrasound and hyperthermia exert a synergistic action. The effect was shown to depend on ultrasound intensity and the temperature at which the treatment takes place. The temperature range enhancing the ultrasound effect shifted forward higher temperature with increasing ultrasound intensity. For every intensity value, an optimal temperature exists at which the synergetic effect is maximum. The biophysical interpretation of the results obtained is based on the assumption that synergism is due to an additional lethal damage, which arises from the interaction of some sub-lesions induced by both agents. These sublesions are considered non-lethal if the agents are applied separately.     

Sodium + potassium-activated ATPase of mammalian brain. Regulation of phosphatase activity.

1. The K+-nitrophenylphosphatase activity associated with mammalian brain (Na+ + K+)-ATPase displays K+ activation curves that have intermediary plateaus and maxima in the presence of less than saturating concentrations of Na+. Zero Na+ and saturating Na+ produce sigmoid K+-activation curves with low and high K+ affinities respectively. 2. ATP inhibits K+-activated nitrophenylphosphatase through both competitive and non-competitive mechanisms. ATP is synergistic with Na+ in the mechanism which converts the enzyme from low to high K+ affinity. 3. The Na+ and K+ interactions can be accounted for by equations which describe a model with separate regulatory sites for Na+ and K+ and with K+- requiring catalytic site which is only accessible in one of the two principal conformational stages of the enzyme. 4. The effects of ATP can be accounted for by the same model through interactions at a single nucleotide binding site. Inhibition which is competitive with K+ and non-competitive with substrate arises from stabilization of the inactive enzyme conformation. Inhibition which is non-competitive with K+ and competitive with substrate results from interactions with the active enzyme conformation. The synergism between Na+ and ATP appears to arise as a consequence of the formation of phosphoryl enzyme. 5. A model for (Na+ + K+)-ATPase is discussed which involves in-phase coupling of subunit interactions as suggested by these studies.     

Copillaria hepatica: Fine structure of egg shell

The structure of the Capillaria hepatica egg shell was studied with the electron microscope and correlated with light microscope histochemical observations. The shell is composed of fibrous and nonfibrous components, both of which stain for protein. The fibrous component, the major portion of the shell, consists of submicroscopic fibers. The nonfibrous component is located in the outer region of the shell but is not always visible; when present it has a reticulated appearance in electron micrographs. The fibrous component is divided into outer and inner regions. The outer region is composed of radially arranged pillars which are connected at their outer surface by a beam-like network and are anchored at the base to a compact inner region. The inner region consists of a series of concentrically arranged lamellae above which is located a nonlaminated region where the pillar bases originate. At each polar end of the shell is a single opening plugged with a material which contains acid mucopolysaccharide. The fine structure of the body of the plug is unresolvable with the electron microscope; its outer surface is impregnated with electron dense particles. Externally the shell is covered by a 250 Å thick continuous membrane which is in close opposition to the surrounding host tissue.     

The ultrastructure of the alimentary tract of the skin-penetrating larva of Nippostrongylus brasiliensis (Nematoda)

The structure of the alimentary tract of the third stage infective larva of Nippostrongylus brasiliensis has been described. The cuticle which lines the buccal cavity and oesophagus differs from that which lines the mouth and covers the external surface of the nematode. The oesophagus is a cellular structure and is not, as previously thought, a syncytium. The secretory granules of the oesophageal glands are surrounded by multi-layered membranes which give a myelinated appearance to the granules. The cells of the oesophago-intestinal junction are lined with cuticle and are presumably part of the stomodaeum. The intestine is thin-walled and the cells bear short, widely spaced microvilli. The lumen of the intestine contains whorls of membranes which are probably phospholipid and could act as a food reserve for the larva. The rectum and anus are lined with cuticle.     

Mechanisms of cell differentiation: Affinity of a morphogenetic factor to DNA

Summary A morphogenetic factor which induces inTriturus gastrula ectoderm tissues which are derived from mesoderm and endoderm has been extracted from chicken and amphibian embryos. The factor which is protein in nature has been obtained from chicken embryos in a highly purified state.The biological activity of the chicken factor is partially inhibited when the factor is combined with chicken DNA or sonicated chicken DNA.When the 3H-labelled factor is combined with sonicated DNA and then centrifuged on a sucrose gradient the factor migrates in part with the DNA. This indicates that the factor is bound to DNA.The inferences from these results are discussed with regard to the possible mechanism of action of the factor and the molecular mechanism of differentiation.     

Clinical Significance and Pathogenesis of Osteoporosis

The development of osteoporosis with advancing age in man is a widespread if not a universal phenomenon. The average loss between youth and old age amounts to about 15% of the skeleton but involves a much larger proportion of trabecular than of cortical bone.The principal clinical manifestation of osteoporosis is fracture, and three osteoporotic fracture syndromes can be defined: the lower forearm fracture, which predominantly affects women between the ages of 50 and 65; the fracture of the proximal femur, which affects both sexes over the age of 70; and the relatively rare vertebral crush fracture syndrome, which may present at any age but is most common in elderly women.The lower forearm fracture rate is inversely related to the mean normal lower forearm x-ray “density” of the wrist, which falls by about 30% in the 15 years following the menopause. This process, which is associated with corresponding trabecular bone loss elsewhere in the skeleton, is associated with a corresponding rise in the fasting urinary calcium excretion. Some degree of negative calcium balance, and consequent bone resorption, probably occurs in everyone during the later part of the night because calcium absorption is completed within about three to five hours of a meal. In postmenopausal women, however, the sensitivity of the bone to parathyroid hormone appears to be increased, and their nocturnal negative calcium balance therefore comes to exceed the positive balance which can be achieved during the waking hours.Femoral neck fractures in old people reflect the further progression of osteoporosis with advancing age since the fracture rate is inversely correlated with the mean thickness of the metacarpal cortex in the normal population. This progressive osteoporosis is associated with and could well result from a steady decline in calcium absorption which is at least partially attributable to vitamin-D deficiency and reversible on vitamin-D treatment.The vertebral crush fracture syndrome represents a severe degree of spinal osteoporosis which may be associated with relatively normal peripheral bones. It probably results from an accelerated negative calcium balance which mobilizes trabecular bone preferentially. Some of the factors which may contribute to this accelerated negative balance have been identified and include a reduced rate of bone turnover, impaired calcium absorption, and low oestrogen activity as judged by vaginal smears, but there may well be others as yet unidentified.     

Observations on the structure of kefir grains and the distribution of the microflora

The mixed flora of yeasts and lactobacilli of kefir is held together in non-dispersible structures which build up into large grains. The fibrillar extracellular material of the matrix in which the microflora is embedded was stained by ruthenium red and periodic acid-thiosemicarbazide silver proteinate, indicating that it was largely composed of carbohydrate. It is suggested that the carbohydrate is of bacterial origin and that this is produced by a population of lactobacilli which resides within the matrix and which separates non-carbohydrate-producing populations of lactobacilli and yeasts so that sheet-like structures are formed which show asymmetry, with yeasts predominating on one side and lactobacilli on the other.     

Estimation of the amount of inbreeding due to subdivision of a population

In a large population which is subdivided into isolated or partially isolated subpopulations polymorphic for a gene locus, there is an excess of homozygotes due to the subdivision. This excess increases with the variance of the gene frequency. The excess can be measured by the “coefficient of inbreeding.” The aim of this paper is to estimate this coefficient, which is a function of various population parameters. We suggest several different estimates, which are the same functional form of unbiased estimates of the population parameters. These estimates are shown to be consistent. They have been compared by numerical methods among themselves and with two other estimates suggested previously.     

The solution structure of the N-terminal domain of riboflavin synthase

The structure of the amino-terminal domain of Escherichia coli riboflavin synthase (RiSy) has been determined by NMR spectroscopy with riboflavin as a bound ligand. RiSy is functional as a 75 kDa homotrimer, each subunit of which consists of two domains which share very similar sequences and structures. The N-terminal domain (RiSy-N; 97 residues) forms a 20 kDa homodimer in solution which binds riboflavin with high affinity. The structure features a six-stranded antiparallel beta-barrel with a Greek-key fold, both ends of which are closed by an alpha-helix. One riboflavin molecule is bound per monomer in a site at one end of the barrel which is comprised of elements of both monomers. The structure and ligand binding are similar to that of the FAD binding domains of ferrodoxin reductase family proteins. The structure provides insights into the structure of the whole enzyme, the organisation of the functional trimer and the mechanism of riboflavin synthesis. C48 from the N-terminal domain is identified as the free cysteine implicated in a nucleophilic role in the synthesis mechanism, while H102 from the C-terminal domains is also likely to play a key role. Both are invariant in all known riboflavin synthase sequences.     

Reversible reaction of cyanate with a reactive sulfhydryl group at the glutamine binding site of carbamyl phosphate synthetase.

Carbamyl phosphate synthetase from Escherichia coli reacts stoichiometrically (one to one) with [14C]cyanate to give a 14C-labeled complex which can be isolated by gel filtration. The formation of the complex is prevented if L-glutamine is present or if the enzyme is first reacted with 2-amino-4-oxo-5-chloropentanoic acid, a chloro ketone analog of glutamine which has been shown to react with a specific SH group in the glutamine binding site. The rate of complex formation is increased by ADP and decreased by ATP and HCO3-. The isolated complex is inactive with respect to glutamine-dependent synthetase activity. However, the reaction of cyanate with the enzyme is reversible. The rate of dissociation of the isolated complex is not affected by pH (over the pH range 6-10), is greatly increased by ATP and HCO3-, and is decreased by ADP. The allosteric effectors ornithine and UMP have no effect on either the rate of formation or the rate of dissociation of the complex; however, the apparent affinity of the enzyme for ATP is decreased by UMP and increased by ornithine. The site of reaction of cyanate with carbamyl phosphate synthetase, which is composed of a light and a heavy subunit, is with an SH group in the light subunit to give an S-carbamylcysteine residue. The binding of L-[14C]glutamine to the enzyme and the inhibition of glutamine-dependent synthetase activity by the chloroketone analog are both prevented by the presence of cyanate. The reaction with cyanate is considered to be with the same essential SH group which is located in the glutamine binding site and is alkylated by 2-amino-4-oxo-5-chloropentanoic acid. The bicarbonate-dependent effects of ATP suggest that formation of the activated carbon dioxide intermediate is accompanied by changes in the heavy subunit which functionally alter the properties of the glutamine binding site on the light subunit. The allosteric effects of ornithine and UMP are probably not related to this intersubunit interaction.     

几种培养方法对萼花臂尾轮虫(Brachionus calyciflorus)种群密度及其环境因子影响的比较研究

分别利用泥鳅 -藻 -轮虫和罗非鱼 -藻 -轮虫共生隔离培养的方法 ,研究萼花臂尾轮虫 ( Brachionuscalyciflorus)的种群密度和主要环境因子的变化。研究结果表明 :加入泥鳅的培养缸中 ,有 1 1 d轮虫数量超过 1 0 0 ind/ml,其中有 5d超过 2 0 0ind/ml,最高密度达 2 50 ind/ml;在罗非鱼的培养缸中 ,在整个实验过程中 ,种群密度超过 1 0 0 ind/ml以上的时间有 6d(第5～ 1 0天 ) ;单纯利用藻类培养轮虫时 ,种群密度较低 ,且维持时间较短 ,最高种群密度不足 1 0 0 ind/ml,利用藻类 -轮虫 -鱼类共存同一生态系统中 ,并使鱼类和轮虫隔离进行生态培养时 ,能提高轮虫种群密度 ,延长种群高峰期     

Analysis of methylthiohydantoins of amino acids by gas-liquid chromatography of their trimethylsilyl derivatives

A procedure is described for the analysis of methylthiohydantoins of amino acids from the Edman degradation of proteins and peptides by gas-liquid chromatography of their trimethylsilyl derivatives. The procedure is applicable to the methylthiohydantoins of all the amino acids commonly found in proteins with the exception of arginine, which did not yield a volatile derivative, and hydroxyproline and hydroxylysine, which were not investigated. Chromatographic separation is achieved in a single run with only one unresolved pair, which can be separated by a supplementary procedure requiring only 4.5 min.     

Synthesis of bioprocesses using physical properties data

The aim of this article is to illustrate and evaluate a synthesis procedure which has been extended to tackle bioprocesses. Physical property information is used to screen candidate units thereby reducing the size of the synthesis problem. In this way, only units which exploit large property differences between components in a stream are selected. This is important for bioprocesses because of the large number of components and wide range of unit operations which are available. The screening technique and bioprocess-unit-design methodologies have been incorporated within an implicit enumeration algorithm which was developed for chemical process synthesis and is implemented in Java programming language. An important advantage is the ability of the bioprocess synthesis software to generate a ranked list of flowsheets which may subsequently be analyzed in more detail. Two case studies are used to evaluate the bioprocess-synthesis technique. The first system involves a product which is secreted from the host organism. The second has significantly different characteristics in that the product is intracellular and forms inclusion bodies. The latter case study, in particular, is a large synthesis problem with 12 unit operations and 20 contaminant compounds. The results show that the synthesis methodology identifies a set of economically optimal flowsheets in a reasonable computational time which demonstrates its ability to deal with large synthesis problems. Using the synthesis methodology we can generate bioprocesses which are optimal in a system-wide, rather than unit-by-unit, sense.     

Prediction and rationalization of the pH dependence of the activity and stability of family 11 xylanases

This paper presents a study of the pH dependence of the activity and stability of a set of family 11 xylanases for which X-ray structures are available, using the PROPKA approach. The xylanases are traditionally divided into basic and acidic xylanases, depending on whether the catalytic acid is hydrogen bonded to an Asn or Asp residue. Using X-ray structures, the predicted pH values of optimal activity of the basic xylanases are in the range of 5.2-6.9, which is in reasonable agreement with the available experimental values of 5-6.5. In the case of acidic xylanases, there are only four X-ray structures available, and using these structures, the predicted pHs of optimal activity are in the range of 4.2-5.0, compared to an observed range of 2-4.6. The influence of dynamical fluctuations of the protein structure is investigated for Bacillus agaradhaerens and Aspergillus kawachii xylanase using molecular dynamics (MD) simulations to provide snapshots from which average values can be computed. This decreases the respective predicted pH optima from 6.2-6.7 and 4.8 to 5.3 +/- 0.3 and 4.0 +/- 0.2, respectively, which are in better agreement with the observed values of 5.6 and 2, respectively. The change is primarily due to structural fluctuations of an Arg residue near the catalytic nucleophile, which lowers its pKa value compared to using the X-ray structure. The MD simulations and some X-ray structures indicate that this Arg residue can form a hydrogen bond to the catalytic base, and it is hypothesized that this hydrogen bond is stabilized by an additional hydrogen bond to another Glu residue present only in acidic xylanases. Formation of such a hydrogen bond is predicted to lower the pH optimum of A. kawachii xylanase to 2.9 +/- 0.3, which is in reasonable agreement with the observed value of 2. The predicted pH of optimal stability is in excellent agreement with the pH value at which the melting temperature (Tm) is greatest. Some correlation is observed between the pH-dependent free energy of unfolding and Tm, suggesting that the thermostability of the xylanases is partly due to a difference in residues with shifted pKa values. Thus, the thermostability of xylanases (and proteins in general) can perhaps be increased by mutations that introduce ionizable residues with pKa values significantly lower than standard values.     

Multiple interactions of a DNA-binding protein in vivo. II. Effects of host mutations on DNA replication of phage T4 gene 32 mutants.

A quantitative analysis has been made of the kinetics of disulphide bond formation, breakage, and rearrangement which occur during the folding and unfolding of the pancreatic trypsin inhibitor. The results have been used to infer the energetics of the protein conformational transitions which accompany each step.The folding transition is shown to be a co-operative process in which all intermediate states with one or two disulphide bonds are unstable relative to the unfolded, reduced protein and that in the fully folded conformation with three disulphide bonds. The approximate two-state nature of the transition at equilibrium was demonstrated experimentally. The folding transition of the inhibitor which has been determined kinetically is therefore analogous to that observed generally with other globular proteins.     

Ultrastructural organization of the ocelloide of Nematodinium. Phylogenetic aspect of the evolution of Warnowiidae Lindemann dinoflagellates photoreceptor (author's transl)

In this study, more details of the different parts of the "ocelloide" of Nematodinium are given. The "hyalosome" which has a radial symmetry is made of endoplasmic reticulum, except the mitochondrial layer. There is only one constrictor ring and only a beginning of the periocellar gallery. The basal sheet on which the crystalline lens lies is a local differentiation of the cortical layer which is observed on the ceiling of the ocellar camera. This camera is related to the intercingular sulcus. There is also an ocellar fiber. The "melanosome" which is made of a double membrane and of many couples of thylakoids is flattened dorso-ventrally. The structure of the retina is similar to that observed in the other Warnowiidae. The pigmented ring is rather well developed and is in continuity with the upper part of the retina. Owing to the simpler organization of its ocelloide, Nematodinium is situated before the genera Warnowia and Erythropsidinium in the evolutionary series.     

Observations on the osmoregulatory system of pentastomids: The tegumental chloride cells

The chitinous procuticle of pentastomids is permeated by small pores which communicate with specialized epidermal cells previously termed cuticular cells. Ultrastructural evidence is presented which suggests that these cells are concerned with the regulation of the hydromineral balance of the haemolymph. Probably all pentastomids regulate hypo-osmotically: that is they maintain a blood concentration which is lower than that of their host. Since pentastomids feed on host blood or lymph which is rich in sodium and chloride, it suggested that chloride cells secrete excess ions. The extent to which chloride cells must concentrate ions depends upon the nature of the food, the rate of exosmosis and the rate of salt gain by diffusion.