The Effects of Brown Rust and Phosphate on Cold Acclimation in Winter Barley

The green leaf area of winter barley, cv. Sonja, sampled fromthe field at different times during winter was always greatestin plants grown at high soil phosphate and smallest in plantsgrown at low soil phosphate, and at each fertilizer level wasgreater in healthy plants than in plants infected by rust (Pucciniahordei). In leaves that survived the coldest period of winter,the percentage area that was damaged was increased by rust infectionwhich prevented the ameliorating effects of high soil P. Rustand low P interacted to reduce the increases in leaf area andshoot d. wt that occurred when higher temperatures prevailedin spring. Under controlled conditions in the laboratory, phosphate reducedthe injury suffered when plants not acclimated to low temperatureswere exposed to freezing conditions, but this effect was removedby rust infection. After rust infection, freezing temperatureswere damaging even to acclimated plants, particularly if grownwith low soil P. Evidence of visible symptoms, and quantitativemeasurements of electrolyte efflux from intact leaves, chlorophyllfluorescence in vivo, and ethane and ethylene evolution fromcold-acclimated plants, showed that infection raised the minimumtemperature at which tissues could survive without injury. Infectedleaves were more sensitive to low temperature post-sporulationthan presporulation. Measurements of electrolyte efflux andchlorophyll fluorescence on plants growing under cold conditionsshowed that infection inhibited the processes of acclimationto low temperatures. Winter barley, Puccinia hordei, injury, low temperature, acclimation     

Electrostatic forces at helix-coil boundaries in DNA

The Tm of internal loop-forming (dA.dT)N domains in pBR322 DNA has been measured over a tenfold range of [Na+]. The slopes SN = dTm/d log [Na+] are linear and decrease in magnitude with decreasing loop size N, signaling a reduction in Na+ released during the transition of these domains to the coil state. Values of SN decrease linearly with increasing N-1 in accordance with the expectation of a simple model for the occurrence of a gradient of long-range electrostatic forces at helix-coil boundaries, and extrapolate almost precisely to the value of S infinity observed for (dA.dT) infinity. These results indicate (1) less counterion is released per phosphate residue from the finite loop than from the infinite-sized loop, and (2) the difference in binding is constant for each boundary formed and independent of the size of the loop within the range examined: approximately 350 base pair (bp) greater than N greater than 71 bp. The slope of the dependence of SN on N-1 indicates the region of higher charge density at the boundary extends at least 18 A into the coil and probably 40-50 A before dropping to a value characteristic of the unperturbed coil. The free energy for excess counterion binding at boundaries can be expressed by -delta G/RT = 10.47 log[Na+] + 5.234 When the loop entropy function in a statistical mechanical algorithm for the dissociation of DNA is weighted by this quantity, calculated Tm are seen to vary by only +/- 0.09 degrees C from observed.     

Recolonisation of four small streams in central Scotland following drought conditions in 1984

Four streams in the Loch Ard Forest in central Scotland dried out almost completely during a drought in the summer of 1984. The recovery of the invertebrate populations in the streams was studied from September 1984 until March 1985 when most of the insect larvae and nymphs were almost full-grown.The appearance of very small larvae belonging to several insect orders within a month of the streams' filling up suggests that they had survived the drought as eggs, or eggs had been laid by adults soon after the drought ended.Statistical analysis showed that with the exception of 3 taxa there was no significant difference between the numbers of animals in November 1984 samples and in samples collected in March 1985.Comparison of samples from March 1984 and March 1985 showed significant difference in population size for a few species; with 2 exceptions the 1984 samples (ie before drought) were larger. In the long term the overall effect of the drought on the invertebrate communities seems to have been limited.     

Evidence for direct arrhythmogenic action of endothelin.

We studied electrophysiological effects of endothelin on canine cardiac tissues. Endothelin prolonged action potential duration and decreased spontaneous firing rate of the right bundle branch cells. At a concentration of 2 x 10(-7)M the plateau phase of action potentials was flattened, followed by the abrupt occurrence of early afterdepolarizations (EADs). ET, at a concentration as low as 2 x 10(-9)M, was capable of inducing EADs although their incidence was low. The EADs were initiated from the membrane potential less negative than -30mV and were suppressed by nicardipine, suggesting the involvement of dihydropyridine-sensitive Ca2+ channels in the induction of EADs. Because EADs are considered to underlie certain types of arrhythmias endothelin per se may have arrhythmogenic action.     

Topological disposition of the sequences -QRKIVE- and -KETYY in native (Na+ + K+)-ATPase

The dispositions with respect to the plane of the membrane of lysine-905 in the internal sequence -EQRKIVE- and of lysine-1012 in the carboxy-terminal sequence -RRPGGWVEKETYY of the alpha-polypeptide of sodium and potassium ion activated adenosinetriphosphatase have been determined. These lysines are found in peptides released from the intact alpha-polypeptide by the extracellular protease from Staphylococcus aureus strain V8 and by trypsin, respectively. Synthetic peptides containing terminal sequences of these were used to prepare polyclonal antibodies, which were then used to prepare immunoadsorbents directed against the respective peptides. Sealed, right-side-out membrane vesicles containing native (Na+ + K+)-ATPase were labeled with pyridoxal phosphate and sodium [3H]borohydride in the absence or presence of saponin. The labeled alpha-polypeptide was isolated from these vesicles and digested with appropriate proteases. The incorporation of radioactivity into the peptides binding to the immunoadsorbent directed against the sequence pyrERXIVE increased 3-fold in the presence of saponin as a result of the increased accessibility of this portion of the protein to the reagent when the vesicles were breached by saponin; hence, this sequence is located on the cytoplasmic face of the membrane. It was inferred that the carboxy-terminal sequence -KETYY is on the extracytoplasmic face since the incorporation of radioactivity into peptides binding to the immunoadsorbent directed against the sequence -ETYY did not change when the vesicles were breached with saponin.     

Inactivation of pyruvate decarboxylase by 3-hydroxypyruvate.

Pyruvate decarboxylase from Zymomonas mobilis is inhibited by 3-hydroxypyruvate, which can also act as a poor substrate. While catalysing the decarboxylation of this alternative substrate, the enzyme undergoes a progressive but partial inactivation over several hours. The extent of inactivation depends upon the pH and upon the concentration of 3-hydroxypyruvate. After partial inactivation and removal of unchanged 3-hydroxypyruvate, enzymic activity recovers slowly. We suggest that inactivation results from accumulation of enzyme-bound glycollaldehyde, which is relatively stable, possibly because it is dehydrated to form an acetyl group.     

Solid-phase synthesis of peptides with C-terminal asparagine or glutamine. An effective, mild procedure based on N alpha-fluorenylmethyloxycarbonyl (Fmoc) protection and side-chain anchoring to a tris(alkoxy)benzylamide (PAL) handle

Attempts to anchor Fmoc-asparagine or glutamine as p-alkoxybenzyl esters for solid-phase peptide synthesis are fraught with difficulties. A convenient and effective method to prepare peptides with C-terminal asparagine or glutamine involves quantitative attachment of N alpha-Fmoc-C alpha-tert.-butyl aspartate or glutamate via the free omega-carboxyl groups to a tris(alkoxy)benzylamino (PAL) support. Chain elongation proceeds normally by standard Fmoc chemistry, and treatment with acid, e.g., CF3COOH--CH2Cl2, 90 min at 25 degrees, releases the desired peptides in greater than 95% yields without side reactions at the C-terminus. Feasibility of the approach has been demonstrated by the syntheses of the C-terminal octapeptide from human proinsulin, H-Leu-Ala-Leu-Glu-Gly-Ser-Leu-Gln-OH, and the serum thymic factor pGlu-Ala-Lys-Ser-Gln-Gly-Gly-Ser-Asn-OH.     

Purification and partial characterization of an iron regulated outer membrane protein of B. fragilis under non-denaturing conditions

The CHAPS-PAGE gelsystem we applied gave a good separation of the proteins of Bacteroides fragilis under non-denaturing conditions. We succeeded with preparative CHAPS-PAGE in purifying an iron regulated outer membrane protein (a 44 kDa polypeptide on SDS-PAGE) of B. fragilis. This integral membrane protein proved to be a lipopolysaccharide binding protein with an isoelectric point of approximately pH 5.5. This method of purifying membrane proteins could be an important step in research into the function of membrane proteins.