Showing papers by "Royal Holloway, University of London published in 1985"

L-Phenylalanine ammonia-lyase from Phaseolus vulgaris. Characterisation and differential induction of multiple forms from elicitor-treated cell suspension cultures.

Abstract: 1 l-Phenylalanine ammonia-lyase (EC 4.3.1.5) has been purified over 200-fold from cell cultures of bean (Phaseolus vulgaris L.) exposed to elicitor heat-released from the cell walls of the phytopathogenic fungus Colletotrichum lindemuthianum. Four forms of the enzyme, with identical Mr but differing apparent pI values of 5.4, 5.2, 5.05 and 4.85, were observed following the final chromatofocussing stage of the purification. 2 A preparation (purified 43-fold by ammonium sulphate precipitation, gel-filtration and ion-exchange chromatography) containing all four forms exhibited apparent negative rate cooperativity with respect to substrates. However, the individual forms displayed normal Michaelis-Menten kinetics, with Km values of 0.077 mM, 0.122 mM, 0.256 mM and 0.302 mM in order of decreasing apparent pI value. 3 A preparation purified 200-fold and containing all four forms was used to immunise rabbits for the production of anti-(phenylalanine ammonia-lyase) serum. The antiserum was characterised by: (a) immunotitration experiments; (b) solid phase enzyme-linked immunosorbent assays; (c) comparison of immunoprecipitates of 35S-labelled phenylalanine ammonia-lyase subunits (synthesized both in vivo and in vitro) on both one-dimensional and two-dimensional polyacrylamide gels after immunoprecipitation with the bean antiserum or antisera raised against pea and parsley phenylalanine ammonia-lyase preparations and (d) immune blotting. 4 Experiments involving (a) immunoprecipitation followed by analysis on SDS/polyacrylamide gels and (b) SDS/polyacrylamide gel electrophoresis followed by immune blotting, indicated that the Mr of newly synthesized (in vivo and in vitro) bean phenylalanine ammonia-lyase subunits is 77000; a 70000-Mr form is readily generated as a partial degradation product during purification. 5 Immunoprecipitates of bean phenylalanine ammonia-lyase synthesized both in vivo and in vitro showed the presence of multiple subunit types of identical Mr but differing in pI. Furthermore, treatment of bean cultures with Colletotrichum elicitor resulted in a 10-fold increase in phenylalanine ammonia-lyase extractable activity within 8 h, and chromatofocussing analysis indicated that this was associated with differential increased appearance of the high-pI, low-Km forms as compared to the two higher Km forms. This differential induction was further confirmed by immune blotting of crude extracts subjected to isoelectric focussing.

Co-ordinated synthesis of phytoalexin biosynthetic enzymes in biologically-stressed cells of bean (Phaseolus vulgaris L.).

Abstract: Changes in the rates of synthesis of three enzymes of phenyl-propanoid biosynthesis in Phaseolus vulgaris L. (dwarf French bean) have been investigated by immunoprecipitation of [35S]methionine-labeled enzyme subunits with mono-specific antisera. Elicitor causes marked, rapid but transient co-ordinated increases in the rate of synthesis of phenyl-alanine ammonia-lyase, chalcone synthase and chalcone isomerase concomitant with the phase of rapid increase in enzyme activity at the onset of accumulation of phenyl-propanoid-derived phytoalexin antibiotics in suspension cultures of P. vulgaris. Co-ordinate induction of enzyme synthesis is also observed in hypocotyl tissue during race:cultivar-specific interactions with Colletotrichum lindemuthianum, causal agent of anthracnose. In an incompatible interaction (host resistant) there are early increases apparently localized to the initial site of infection prior to the onset of phytoalexin accumulation and expression of hypersensitive resistance. In contrast, in a compatible interaction (host susceptible) there is no induction of synthesis in the early stages of infection, but a delayed widespread response at the onset of lesion formation associated with attempted lesion limitation. It is concluded that expression of the phytoalexin defense response in biologically stressed cells of P. vulgaris characteristically involves co-ordinate induction of synthesis of phytoalexin biosynthetic enzymes.

Ikaite, the parent mineral of jarrowite-type pseudomorphs

Abstract: The pseudomorphs known as jarrowite which were found during the middle part of the last century in muds at Jarrow Slake on the River Tyne are now identified as being pseudomorphs after crystals of the mineral ikaite. Ikaite, CaCO 3 .6H 2 O, is a metastable mineral in sedimentary rocks. It crystallizes at near to zero Celsius, but breaks down at normal temperatures to form calcium carbonate and water. Jarrowites are pseudomorphed in calcite which appears to have been generated by redistribution of the calcium carbonate formerly present in the parent ikaite. The high porosity of the jarrowites is the result of the decrease in volume which accompanied the change. The paper reviews the known occurrences of pseudomorphs of jarrowite-type (pseudogaylussite, thinolite, White-Sea hornlets, glendonite, gersternkorner, gennoishi, fundylite and others), the discovery of ikaite and the ways in which the pseudomorphs alter ikaite may have a role as palaeothermometers.

A simulation model for bed allocation to hospital inpatient departments

Abstract: A general simulation model can be used to determine the bed complement in hospital inpatient departments to meet a pre determined demand for service. Certain operational constraints were considered in this version of our model: (1) emergency pa tients should be admitted without delay, (2) occupancy should not fall below a prespecified level, and (3) the waiting list length should not exceed a predetermined number. Extensions and modifications of the model are briefly discussed. Three case studies on the general medical, general surgical and orthopaedics departments at St. Peter's Hospital, Chertsey, Surrey (England), illustrate how this model may be utilised.

Metabolic changes in elicitor-treated bean cells. Selectivity of enzyme induction in relation to phytoalexin accumulation.

Abstract: 1 Treatment of cell suspension cultures of bean (Phaseolus vulgaris cv Immuna) with an elicitor preparation heat-released from the cell walls of the phytopathogenic fungus Colletotrichum lindemuthianum resulted in rapid changes in the composition of the bean cell walls These consisted of (a) increases in phenolic material bound to the cellulosic and hemicellulosic fractions of the wall, (b) loss of material (mainly glucose) from the hemicellulosic fraction and (c) an increase in wall-associated hydroxyproline 2 The increases in wall-bound phenolics were preceded by (a) rapid decreases in the intracellular levels of free hydroxycinnamic acids and (b) transient increases in the extractable activities of l-phenylalanine ammonia-lyase and cinnamic acid 4-hydroxylase 4-Hydroxycinnamic acid 3-hydroxylase activity was present at a high level in control cultures and was not induced by elicitor Changes in the levels of cytochrome P-450, as determined by dot blot assays utilising an anti-(P-450) monoclonal antibody, paralleled the changes in cinnamic acid 4-hydroxylase activity 3 The accumulation of cell wall hydroxyproline was associated with rapid transient increases in the extractable activities of proline 2-oxoglutarate dioxygenase and a protein arabinosyl transferase An hydroxyproline-rich acceptor protein of Mr 42 500 was the major protein to incorporate [3H]arabinose following elicitation of the bean cells, and the kinetics of the extent of labelling of this protein paralleled the accumulation of hydroxyproline protein in the endomembrane system 4 The above metabolic changes associated with cell wall components followed rapid kinetics similar to those involved in the formation of the phytoalexin kievitone in the elicited cultures [Robbins, M P et al (1985) Eur J Biochem 148, 563–569] It is therefore concluded that increased 5-hydroxy-substituted isoflavonoid biosynthesis, wall-bound phenolic synthesis and synthesis of arabinosylated hydroxyproline-rich protein are all early events which are closely linked to the initial interaction between plant cell and fungal elicitor

Homologies of Molar Cusps and Crests, and Their Bearing on Assessments of Rodent Phylogeny

Abstract: Evidence for phylogeny derives basically from morphological comparisons, whether of the structure of protein molecules, chromosomes, skulls, or, in the case of paleontological material, mainly teeth. The concept of homology arises from the recognition that some resemblances are more significant than others that are superficial or accidental. It is necessary to decide, when comparing two species, which structural element in one species should be compared with an element in the other. Such corresponding structures in different animals are given the same name (Gr. homos, same; logos, word), and resemblances and differences between them form the raw data on which phylogenetic hypotheses are based. Whether resemblances are due to inheritance from a common ancestor, or whether they have been produced by parallel evolution, is a question that can be decided only after considerable investigation, if at all. Therefore, in this paper I will use the term homology in its etymological sense, to mean morphologically comparable structures, whether or not their resemblance is due to common ancestry.

Ancient archives: the edict of Alexander to Priene, a reappraisal

Abstract: ‘The historical value of an object depends not so much on the nature of the object as on its associations, which only scientific excavation can detect.’ The full significance of an inscription may equally rely on knowledge of its archaeological context. In practice, however, users of inscriptions often neglect this aspect. The standard commentaries, new and old, on Alexander's famous ‘edict to Priene’ (hereafter ‘AE’) tend to ignore the physical context of the inscription (I.Priene I) and to treat the text as an isolated or one-off document. Consequently no-one reading Dittenberger, Tod or now Heisserer would learn that it is one of a series of public inscriptions with a consistent theme belonging to an ‘archive’ of connected texts. The inscription is not discussed as one of a group of documents, its monumental setting is largely ignored and the rich corpus of Prienian inscriptions is not exploited fully as a control and source for the historical background of the AE. It is the purpose of this article to try to show that the AE cannot be properly studied in this archaeological limbo. ‘The associations’ of the AE are vital. They provide a new perspective from which to study the text.

The in Vitro Biosynthesis of Carotenoids

Abstract: Publisher Summary This chapter presents the enzymology of carotenoid biosynthesis in the light of in vitro studies. Despite the increase over recent years in the number of cell-free systems capable of in vitro carotenoid biosynthesis, understanding of the enzymology of carotenoid formation is still in its infancy. The carotenoid enzymologist faces a number of practical difficulties. The modern approaches to the purification of enzymes, using techniques such as affinity chromatography, chromatofocusing, and fast protein liquid chromatography, have the considerable advantages of being rapid and also of being applicable to relatively crude protein preparations. Cell-free preparations from organisms such as A. carterae, Aphanocapsa, Flavobacterium, Halobacterium, and Phycomyces look very promising as systems for the purification of carotenogenic enzymes.

Allocating Doctors to Shifts in an Accident and Emergency Department

Abstract: A method is proposed for allocating doctors to weekly shifts in an accident and emergency department of a hospital. Two models, solved by dynamic programming, are used. The first finds the optimal solution to the problem of allocating doctors per hour of the week proportionally to the corresponding patient arrival-rate; and the second uses this solution to smooth out the hourly discrepancies in the number of doctors within a shift. The solution is then assessed by computer simulation.

Structure and function in the excretory system of archaeogastropods and their significance in the evolution of gastropods

Abstract: Three species of archaeogastropod mollusc, Monodonta lineata (da Costa), Emarginula reticulata Sowerby and Patella vulgata L. were selected as representative members of the Trochacea, Fissurellacea and Patellacea, respectively, for a comparative anatomical and ultrastructural study of the excretory system. Primary urine formation takes place by filtration of blood through the walls of the paired auricles in Monodonta and Emarginula and of the single auricle and ventricle in Patella. Urine then passes to right and left kidneys along the renopericardial canals. Contrary to earlier reports the two kidneys are different in structure and function in all three species, the larger right kidney retaining the primitive function of nitrogenous excretion, the left having a predominantly resorptive role and with a capacity to abstract from the blood solutes of larger molecular mass. The difference in the size of the two kidneys is exaggerated in Patella and Emarginula as a consequence of partial restoration of bilateral symmetry in these limpets. It has been possible to demonstrate at the ultrastructural level that the minute left kidney of Emarginula is functional. The vacuolated epithelial cells of the right kidney contain layered excretory spherules composed of purines, melanin and ferric iron in different proportions in the three genera. There is close similarity in the ultrastructural organization of these cells in Monodonta and Emarginula, but those of Patella show marked differences and their excretory spherules contain a higher proportion of melanin. The position of the left kidney in the mantle skirt, as exemplified by Monodonta, is believed to have arisen in the earliest gastropods correlated with the development of helical coiling. This was accompanied by a change in its blood vessels. It has lost its afferent renal vein, which primitively would have carried deoxygenated blood from the viscera, an arrangement which persists in the right kidney. The left efferent renal vein is reduced in Monodonta and lost in Patella and Emarginula. A new vessel has arisen linking left auricle and left kidney and there is evidence to suggest that it carries post-branchial oxygenated blood. It is believed to serve as both an afferent and major efferent route. The physiological implications of this change in the blood supply are discussed and held to be responsible for the functional differences between the two kidneys, creating conditions in the left which favour resorption of organic solutes and ions, and leaving the right kidney with the primary role of nitrogenous excretion. The evolution of the nephridial gland is examined in this context and is also believed to be correlated with the change in the blood supply to the left kidney. Ultrastructural evidence is given in support of its suggested resorptive function. The significance of the differences between right and left kidneys of archaeogastropods is discussed in relation to the evolution of the monotocardian excretory system, and the possible phylogenetic relationships of the groups of archaeogastropods are considered.

Carotenoid biosynthesis by Aphanocapsa homogenates coupled to a phytoene-generating system from Phycomyces blakesleeanus

Abstract: The ability of a cell extract from the C5 carB10(-) (phytoene-accumulating) strain of Phycomyces blakesleeanus to convert [2-14C]mevalonic acid into phytoene has been exploited in a coupled assay with a cell homogenate from the cyanobacterium Aphanocapsa 6714 (Pasteur Culture Collection]. Incubations containing both cell extracts convert [2-14C]mevalonic acid into unsaturated carotenes and xanthophylls of Aphanocapsa, via the [14C]phytoene formed by the C5 preparation. The major advantage of this experimental approach is that it simplifies and shortens the assay procedure, since it eliminates the necessity to initially prepare [14C]phytoene and then add it to the incubation with an emulsifying agent.

Radial histograms as an aid in the inspection of circular objects

Abstract: This paper considers the inspection of circular objects. Template matching forms a competitive approach for inspecting these products, but radial histograms are found to be especially useful whenever products are nearly symmetric. Various methods of renormalising radial histograms are compared, with particular reference to accuracy and resolution. It is shown that radial histogramming is valuable for checking quickly for various defects, and that it can lead to increased accuracy of radial measurement. Its capabilities depend strongly on the previous accurate location of the centres of circular regions: the Hough transform approach is well adapted to this purpose.

Carotenoids and bile pigments in Danaid and swallowtail butterflies

Abstract: The carotenoids found in adult kite swallowtails (Graphium spp.) and the Danaid glassy tigers (Ideopsis, Tirumala and Parantica), butterflies with exposed blue/green pigmented wing membranes (which contain pterobilins) and a modified scale vestiture, were compared with those of related species of Papilios and Danaids with concealed wing membranes an an entire scale vestiture. It was found that both the kite swallowtails contained only lutein, and even lacked β-carotene, the first known instance among butterflies. This was presumably due to selective storage by the larva since the food-plant was found to be rich in varied carotenoids. A possible link between the high concentration of pterobilins in the wing membrane and the lack of carotenoids is discussed.

Shear thinning of the Lennard-Jones fluid by molecular dynamics

Abstract: Extensive Molecular Dynamics, MD, calculations of the Lennard-Jones, LJ, rheological equation of state have been made. Non-equilibrium MD permits evaluation of shear thinning of the dense LJ liquid which adheres in behaviour quite closely with that of more complex “real molecules”. However, quantitative correspondence with simple analytic formulae for non-Newtonian behaviour used in the treatment of experimental data is hindered by poor prediction of certain key parameters. For example, at low shear rates, the equilibrium Newtonian viscosity and, at high shear rates, a limiting shear stress are often required. Both are difficult to obtain by simulation in the portion of the LJ phase diagram which exhibits significant shear thinning and using present techniques. Suggestions for improving the Eyring model for shear thinning are made.

The Chromosomes of Dinoflagellates

Abstract: Publisher Summary The chromosomes of dinoflagellates have long been suspected of being unusual and, in the larger species at least, can be seen even without any special staining. This chapter provides an overview of the historical work and makes an attempt to bring together a current view on the structure of the chromosome. Structurally, dinoflagellates probably have more in common with the nuclei of bacteria, but in terms of their behavior, they show similarities with normal eukaryotic chromosomes; therefore, little is known about their genetic capabilities that it is impossible at this stage to make postulates about their affinities in this respect. While there is very much to be discovered about these structures, it seems clear that they provide either a fascinating bridge between the nuclear arrangements in prokaryotes and those in eukaryotes or a quite remarkable and successful experiment, which, like the dinosaurs, had only limited evolutionary potential.