Function of an internal bacteriophage T7 core during assembly of a T7 procapsid.

A DNA-free, proteinaceous procapsid of bacteriophage T7 (capsid I) has been shown in previous studies to consist of an external, spherical shell (envelope) and an internal, cylindrical core with fibrous projections that connect the core to the envelope. To determine the role of the core in assembly of the envelope of capsid I, the kinetics of appearance of capsid I and possible intermediates in capsid I assembly (AG particles) were determined in the presence and absence of the core. For obtaining these data, agarose gel electrophoresis was used and appeared to be a technique more accurate and efficient than techniques used for obtaining similar data in the past. The results of these experiments were: (i) in the presence of the core, AG particles behaved kinetically as intermediates in the assembly of capsid I; (ii) in the absence of the core, assembly of capsid I terminated prematurely and AG particles accumulated. These and other data have been interpreted by assuming that: AG particles are breakdown products of precursors of capsid I; these precursors have uncorrected errors in the assembly of their envelope; and a function of the core is to correct these errors.     

Isolation of an Hfr donor of Pseudomonas aeruginosa PAO by insertion of the plasmid RP1 into the tryptophan synthase gene

Summary A derivative of the IncP-1 plasmid RP1, temperature-sensitive for maintenance, was inserted into the Pseudomonas aeruginosa chromosome by selection for a plasmid marker (carbenicillin resistance) at nonppermissive temperature. In one strain, PAO 1000, the plasmid was stably integrated in the trpA, B gene cluster mapped at 27 min, as shown by the following evidence. (i) Trp⁺ transductants lost all plasmid markers. (ii) Cleared lysates of PAO 1000 showed no plasmid band typical of the autonomous RP1 in agarose gel electrophoresis. (iii) No transfer of carbenicillin resistance by PAO 1000 was detectable. (iv) PAO 1000 mobilised the chromosome from an origin at, or very near, the plasmid insertion site with high frequency (recovery of proximal markers 10^–3 per donor). Matings on the plate with and without interruption of conjugation showed that chromosome transfer was unidirectional. (v) Recombinants from PAO 1000-mediated crosses did not inherit plasmid markers or the trpA, B mutation. A derivative of PAO 1000 was obtained which had lost the Hfr property and all plasmid markers except carbenicillin resistance. This strain (PAO 1001), when carrying the autonomous RP1 plasmid, was capable of unidirectional chromosome mobilisation like PAO 1000, but with 50-fold lower efficiency. We propose that integration of the temperature-sensitive RP1 plasmid in PAO 1000 occurred via transposition of Tnl, the element specifying carbenicillin resistance.     

Systematic variation in amino acid compositions of grass caryopses

Variation in amino acid patterns of 121 species (72 genera) of grass caryopses is extensively consistent with taxonomic groupings. The patterns of pooids and chloridoids are distinguishable from one another and from those of eu-panicoids and andropogonoids; the bamboos, Oryza, Stipeae, Ehrharta and Microlaena, which share certain morphological and anatomical features, also share a characteristic amino acid profile, while profiles of danthonoioids, Triodia and Aristida are clearly non-pooid. Caryopsis amino acid patterns vary independently of photosynthetic pathway. Embryos from taxonomically diverse genera all show very similar amino acid profiles, which differ strikingly from those of the endosperms, and the amino acid patterns of whole caryopses are dominated by their endosperms, which are responsible for the taxonomic variation. ‘Chemical scores’ of the caryopsis proteins, but not total protein contents, correlate to some extent with taxonomic groupings.     

Variations in Kinetic Properties of Ribulose-1,5-bisphosphate Carboxylases among Plants

Studies of ribulose-1,5-bisphosphate (RuBP) carboxylase from taxonomically diverse plants show that the enzyme from C(3) and crassulacean acid metabolism pathway species exhibits lower K(m)(CO(2)) values (12-25 micromolar) than does that from C(4) species (28-34 micromolar). RuBP carboxylase from aquatic angiosperms, an aquatic bryophyte, fresh water and marine algae has yielded consistently high K(m)(CO(2)) values (30-70 micromolar), similar in range to that of the enzyme from C(4) terrestrial plants. This variation in K(m)(CO(2)) is discussed in relation to the correlation between the existence of CO(2)-concentrating mechanisms for photosynthesis and the affinity of the enzyme for CO(2). The K(m)(RuBP) of the enzyme from various sources ranges from 10 to 136 micromolar; mean +/- sd = 36 +/- 20 micromolar. This variation in K(m)(RuBP) does not correlate with different photosynthetic pathways, but shows taxonomic patterns. Among the dicotyledons, the enzyme from crassinucellate species exhibits lower K(m)(RuBP) (18 +/- 4 micromolar) than does that from tenuinucellate species (25 +/- 7 micromolar). Among the Poaceae, RuBP carboxylase from Triticeae, chloridoids, andropogonoids, Microlaena, and Tetrarrhena has yielded lower K(m)(RuBP) values (29 +/- 11 micromolar) than has that from other members of the grass family (46 +/- 10 micromolar).     

Unsaturated fatty acid but not ergosterol is essential for high ethanol production in Saccharomyces

Summary The membrane lipid composition of Saccharomyces was manipulated by growing cells anaerobically with or without ergosterol and unsaturated fatty acid. Cells low in ergosterol but enriched in unsaturated fatty acid residues on membrane phospholipids produced high concentrations, 13–15.5% w/v, of ethanol at substrate conversion efficiencies of around 90%.     

Measurement of surface potential and surface charge densities of sarcoplasmic reticulum membranes

Summary The binding of the anionic fluorescent probe 1-anilino-8-naphthalene-sulfonate (ANS^–) was used to estimate the surface potential of fragmented sarcoplasmic reticulum (SR) derived from rabbit skeletal muscle. The method is based on the observation that ANS^– is an obligatory anion whose equilibrium constant for binding membranes is proportional to the electrostatic function of membrane surface potential, exp(e₀/kT, where ₀ is the membrane surface potential,e is the electronic charge, andkT has its usual meaning. The potential measured is characteristic of the ANS^– bindings of phosphatidylcholine head groups and is about one-third as large as the average surface potential predicted by the Gouy-Chapman theory. At physiological ionic strength the surface potentials, measured by ANS^–, referred to as the aqueous phase bathing the surface, were in the range –10 to –15 mV. This was observed for the outside and inside surfaces of the Ca²⁺-ATPase-rich fraction of theSR and for both surfaces of theSR fraction rich in acidic Ca²⁺ binding proteins. The inside and outside surfaces were differentiated on the basis of ANS^– binding kinetics observed in stopped-flow rapid mixing experiments. A mechanism by which changes in Ca²⁺ concentration could give rise to an electrostatic potential across the membrane and possibly result in changes in Ca²⁺ permeability.The dependence of the surface potential on the monovalent ion concentration in the medium was used together with the Gouy-Chapman theory to determine the lower limits for the surface charge density for the inside and outside surfaces of the two types ofSR. Values for the Ca²⁺-ATPase richSR fraction were between 2.9×10³ and 3.8×10³ esu/cm², (0.96×10^–6 and 1.26×10^–6 C/cm²) with no appreciable transmembrane asymmetry. A small amount of asymmetry was observed in the values for the inside and outside surfaces of the fraction rich in acidic binding proteins which were ca. 6.6×10³ and ca. 2.2×10³ esu/cm² (2.2×10^–6 and 0.73×10^–6 C/cm). The values could be accounted for by the known composition of negatively-charged phospholipids in theSR. The acidic Ca²⁺ binding proteins were shown to make at most a small contribution to the surface charge, indicating that their charge must be located at least several tens of Å from the membrane surface. The experiments gave evidence for a Donnan effect on the K⁺ distribution in the fraction rich in acidic binding proteins. This could be accounted for by the known concentration of acidic binding proteins in thisSR fraction.The equilibrium constant for ANS^– was shown to be more sensitive to changes in the divalent cation concentration than to changes in the monovalent cation concentration, as predicted by the Gouy-Chapman theory. Use of these findings together with the stopped-flow rapid mixing techniques constitutes a method for rapid and continuous monitoring of changes in ion concentrations in theSR lumen.     

Application of pattern recognition and feature extraction techniques to volatile constituent metabolic profiles obtained by capillary gas chromatography

The applicability of threshold logic units, a form of nonparametric pattern recognition, to the processing of metabolic profile data obtained by high-efficiency glass capillary column gas chromatography has been investigated. The test data included profiles of the volatile constituents of urine from normal individuals and from individuals with diabetes mellitus. A feature extraction algorithm allowed for dimensionality reduction and indicated the constituents most important in the normal versus pathological distinction. With an optimum number of dimensions, a normal versus pathological prediction rate of 93.75% was achieved. Gas chromatography—mass spectrometry was utilized to identify important profile constituents.