Control of activity states of heart mitochondrial ATPase. Role of the proton-motive force and Ca2+

The ATPase complex of submitochondrial particles exhibits activity transitions that are controlled by the natural ATPase inhibitor (Gómez-Puyou, A., Tuena de Gómez-Puyou, M. and Ernster, L. (1979) Biochim. Biophys. Acta 547, 252–257). The ATPase of intact heart mitochondria also shows reversible activity transitions; the activation reaction is induced by the establishment of electrochemical gradients, whilst the inactivation reaction is driven by collapse of the gradient. In addition it has been observed that the influx of Ca²⁺ into the mitochondria induces a rapid inactivation of the ATPase; this could be due to the transient collapse of the membrane potential in addition to a favorable effect of Ca²⁺-ATP on the association of the ATPase inhibitor peptide to F₁-ATPase. This action of Ca²⁺ may explain why mitochondria utilize respiratory energy for the transport of Ca²⁺ in preference to phosphorylation. It is concluded that the mitochondrial ATPase inhibitor protein may exert a fundamental regulatory function in the utilization of electrochemical gradients.     

Cell signaling and heat shock protein expression

Exposure of cells and organs to heat shock is associated with numerous changes in various cellular metabolic parameters and overexpression of proteins collectively known as heat shock proteins (HSP). In this communication we review the cell-signaling events that are altered in response to heat shock as they relate to the subsequent induction of HSP 70 kd (HSP-70) expression. We also review the mechanisms by which HSP-70 is involved in conferring cytoprotective effects. The possibility of altering HSP expression through manipulations of the cell-signal process has clinical importance.The opinions or assertions contained herein are the private views of the authors and are not to be construed as official or reflecting the views of the Department of the Army or Department of Defense.     

A tetrameric allyl complex of sodium, and computational modeling of the Na-allyl chemical shift

Transmetallation of Li[A′] (A′ = [1,3-(SiMe₃)₂C₃H₃]⁻) with sodium tert-butoxide produces the corresponding sodium salt, which crystallizes from THF/toluene in the form of a cyclic tetramer, {Na[A′](thf)}₄. The Na atoms are in a square planar arrangement, bridged with π-bound allyl ligands; the Na-C distances range from 2.591(3)-2.896(3) Å, with an average of 2.70 Å. The geometries of several model organosodium complexes containing cyclopentadienyl and allyl ligands were optimized with density functional theory methods. The optimized structures were used with the gauge-including atomic orbital (GIAO) method to calculate their ²³Na NMR magnetic shielding values. Unlike the case with NaCp, the chemical shift of unsubstituted Na(C₃H₅) is very sensitive to the presence of coordinated THF (causing a 20 ppm upfield shift); silyl substitution has an even larger effect (30 ppm upfield shift). The observed ²³Na shift of δ −3.3 ppm for Na[A′] in THF-d₈, however, cannot be reliably distinguished from that calculated for the [Na(thf)₄]⁺ cation alone.     

Length–weight relationships of seven fish species in the Yangtze River Estuary and its adjacent waters,Eastern China

The length–weight relationships (LWRs) for seven fish species belonging to five families in the Yangtze River Estuary and its adjacent waters are presented. Cynoglossus gracilis, Cynoglossus macrolepidotus, Ctenotrypauchen chinensis, Collichthys niveatus, Ophichthus apicalis and Erisphex potti were collected monthly in 2009 using trammel nets (the size of smaller mesh net was 1.5 m × 15 m × 4 panels with mesh sizes of 2.5, 3.4, 4.3 and 5.8 cm; the size of larger-mesh net was 2.4 m × 30 m × 4 panels with mesh sizes of 5.0, 6.0, 7.0 and 8.0 cm; soaking overnight). Lophiogobius ocellicauda were collected December in 2020 using shrimp trawl nets (mesh size 2.0 cm). The precision of measurement for the fish specimens is 0.1 cm total length and 0.1 g total weight. The estimated ranges of the parameters a and b for the seven fish species were from 0.0001 to 0.0289 and 2.718 to 3.541, respectively. Two new maximum total length were recorded for Ctenotrypauchen chinensis and Ophichthus apicalis.     

Optimization of ascorbic and uric acid separation in human plasma by free zone capillary electrophoresis ultraviolet detection

In this paper we propose a new fast free zone capillary electrophoresis method for the simultaneous determination of ascorbic acid (AA) and uric acid (UA) in human plasma. We investigated the effect of analytical parameters, such as concentration and pH of borate running buffer, cartridge temperature, and sample treatment, on resolution, migration times, corrected peak areas, and efficiency. A good separation was achieved using a 60.2-cmx75-microm uncoated silica capillary and 100 mmol/L sodium borate buffer, pH 8, when metaphosphoric acid was employed as protein precipitant, in less than 4 min. These conditions gave a good reproducibility of migration times (CV 0.35 and 0.34%) and peak areas (CV 3.2 and 3.1%) for ascorbate and urate, respectively. The limit of detection was 0.5mg/L for both analytes when the detection was performed at 254 nm for AA and at 292 nm for UA. We compared the present method with a validated capillary electrophoresis assay by measuring plasma urate and ascorbate in 32 normal subjects and the obtained data were analyzed by the Passing and Bablok regression.     

Prp5 bridges U1 and U2 snRNPs and enables stable U2 snRNP association with intron RNA

Communication between U1 and U2 snRNPs is critical during pre-spliceosome assembly; yet, direct connections have not been observed. To investigate this assembly step, we focused on Prp5, an RNA-dependent ATPase of the DExD/H family. We identified homologs of Saccharomyces cerevisiae Prp5 in humans (hPrp5) and Schizosaccharomyces pombe (SpPrp5), and investigated their interactions and function. Depletion and reconstitution of SpPrp5 from extracts demonstrate that ATP binding and hydrolysis by Prp5 are required for pre-spliceosome complex A formation. hPrp5 and SpPrp5 are each physically associated with both U1 and U2 snRNPs; Prp5 contains distinct U1- and U2-interacting domains that are required for pre-spliceosome assembly; and, we observe a Prp5-associated U1/U2 complex in S. pombe. Together, these data are consistent with Prp5 being a bridge between U1 and U2 snRNPs at the time of pre-spliceosome formation.     

Evaluation of on-farm veal calves’ responses to unfamiliar humans and potential influencing factors

The human–animal relationship is an important component of the welfare of farm animals and for this reason animal responsiveness tests to humans are included in on-farm welfare assessment schemes that provide indicators for this. However, apart from the behaviour of stockpersons towards their animals, other factors may also influence animals’ reactivity to humans as observed through behavioural tests, which can add a further layer of complexity to the interpretation of test results. Knowledge of these factors may help a better interpretation of differences from one farm to another in the outcome of human–animal relationship tests, and may provide clues for improving the relationship between animals and humans. The main objective of this study was to identify whether management or environmental factors could influence the outcome of human–animal relationship tests in veal calves. Two tests were performed when calves were aged 14.9 ± 1.6 (SD) weeks in 148 veal farms: the voluntary approach of an unfamiliar human standing at the feeding fence and the reaction towards an unfamiliar human who entered the home pen and tried to touch each calf in a standardised way (Calf Escape Test (CET) – score 0 to 4). Questionnaires were filled in and interviews with the stockpersons were performed in order to obtain information on stockpersons, management, animal and building characteristics. The latency to touch an unfamiliar human at the feeding fence was significantly correlated with the CET scores. Total number of calves on the farm, space allowance, breed, environmental enrichment, stockperson's experience and season of observation influenced the percentage of calves that scored 0 in CET (i.e. calves that could not be approached). Type of milk distribution, type of breed and number of calves per stockperson influenced the percentage of calves that scored 4 in CET (i.e. calves could be touched). For both CET0 and CET4, the level of self-reported contacts by the stockperson (analysed only on the French subset of 36 farms) did not influence the results. This paper concludes that according to the tests conducted on veal calves on commercial farms, factors such as milk distribution method, breed of the calves or the level of experience of stockpersons with veal farming can have an impact on the results of tests focusing on human–animal relationships.     

Host recognition by the egg–larval parasitoid Chelonus inanitus: effects of physical and chemical cues

Parasitoid wasps are widely used as biocontrol agents. For successful deployment, it is important to know how they search for and recognize their hosts. They often use chemical cues in host searching and recognition. Little attention has been paid to the use of physical cues, particularly when combined with chemical cues, though a combination of cues may improve searching efficiency. Chelonus inanitus L. (Hymenoptera: Braconidae) is a solitary, egg–larval parasitoid wasp that parasitizes various lepidopteran species, including serious crop pests. In this study, effects of physical and/or chemical cues on host recognition were investigated in C. inanitus by observing its antennal searching, ovipositor probing, and the duration of arrestment. In a physical cue test, egg-mass models with glass beads differing in size were used, and a single-egg model with a specific size of bead. Egg-mass models with beads of 0.4–1.0 mm in diameter arrested C. inanitus, in accordance with egg size and arrangement of the host. In a chemical cue test, extract from egg masses of the host Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae), equivalent to half of a host egg mass and extract diluted 100-fold were used, and the undiluted extract stimulated C. inanitus. Untreated egg-mass models (0.2-mm-diameter glass beads) and diluted egg extract arrested C. inanitus less effectively; however, when glass beads were treated with the diluted extract, arrestment increased to the same degree as the undiluted extract. These results suggest that C. inanitus uses both physical and chemical cues to recognize eggs of its host, S. litura, and that these stimuli act synergistically.     

Roles of Long-range Electrostatic Domain Interactions and K+ in Phosphoenzyme Transition of Ca2+-ATPase

Sarcoplasmic reticulum Ca²⁺-ATPase couples the motions and rearrangements of three cytoplasmic domains (A, P, and N) with Ca²⁺ transport. We explored the role of electrostatic force in the domain dynamics in a rate-limiting phosphoenzyme (EP) transition by a systematic approach combining electrostatic screening with salts, computer analysis of electric fields in crystal structures, and mutations. Low KCl concentration activated and increasing salt above 0.1 m inhibited the EP transition. A plot of the logarithm of the transition rate versus the square of the mean activity coefficient of the protein gave a linear relationship allowing division of the activation energy into an electrostatic component and a non-electrostatic component in which the screenable electrostatic forces are shielded by salt. Results show that the structural change in the transition is sterically restricted, but that strong electrostatic forces, when K⁺ is specifically bound at the P domain, come into play to accelerate the reaction. Electric field analysis revealed long-range electrostatic interactions between the N and P domains around their hinge. Mutations of the residues directly involved and other charged residues at the hinge disrupted in parallel the electric field and the structural transition. Favorable electrostatics evidently provides a low energy path for the critical N domain motion toward the P domain, overcoming steric restriction. The systematic approach employed here is, in general, a powerful tool for understanding the structural mechanisms of enzymes.     

Flow injection–chemiluminescence determination of acyclovir

The chemiluminescence (CL) reaction of acyclovir (ACV)–potassium permanganate, with formaldehyde as an enhancer, was investigated by the flow‐injection system, and a new method is reported for the determination of ACV on the basis of the reaction. The method is rapid, effective and simple for the determination of acyclovir in the range 0.2–80 mg/L, with a limit of detection of 0.06 mg/L (3 S:N), a relative standard deviation (RSD) of 3.7% for the determination of 1.0 mg/L acyclovir solution in 11 repeated measurements. The method has been applied to the determination of acyclovir in pharmaceuticals, with satisfactory results. The possible reaction mechanism is also discussed briefly. Copyright © 2011 John Wiley & Sons, Ltd.