Upstream-downstream movements of aquatic invertebrates in a Rocky Mountain stream

Simultaneous collections of drift and organisms moving either upstream or downstream in association with the substrate were made using a specially designed sampler. Samples were taken in a diel series along a transect across the study riffle of a Colorado foothills stream on six dates over an annual cycle. In addition to longitudinal movements, taxonomic composition and diel periodicity were evaluated. The insect-dominated fauna showed a net downstream displacement. Only the caddisflies Helicopsyche borealis and Hesperophylax occidentalis exhibited net upstream movement, primarily a result of low drift frequencies. The taxonomic composition of moving invertebrates differed from that of the benthos. Drift resembled downstream moving substrate-associated invertebrates in composition, but differed from that of the upstream directed fauna. Taxa collectively exhibited four types of diel patterns: 1) similar downstream (drift and substrate-associated movements) patterns, which generally differed from the upstream pattern; 2) similar benthic (upstream and downstream) patterns, which differed from that of drift; 3) aperiodic patterns; and 4) independent patterns for each type of directional movement. Analysis of size classes based on head capsule width for the mayfly Baetis tricaudatus showed significantly smaller size in stationary individuals compared with moving individuals in the population and revealed that nymphs moving during the day were smaller than those moving at night.     

Immunoprotection of guinea pigs against experimental acute pyelonephritis after the administration of Escherichia coli purified fimbriae

Experimental results of the present study evidenced the following aspects: a) the antigen, prepared from type I Escherichia coli purified fimbriae (H. 2946 strain), induced immunity at urinary tract level; b) the immunoprotection induced by oral vaccination with multiple doses of fimbriated antigen produced a significant decrease of acute pyelonephritis in newborn guinea pigs and at the same time, a local protection of the urinary tract; c) the immunoprophylaxis by vaccine prepared from fimbriae represents a preferential solution for urinary tract infections prevention in general and especially in children; d) the frequency distribution differences between "protected" and "non-protected" animals were evaluated by chi-square--test with YATES correction and proved to be statistically significant at probability levels.     

The complete amino-acid sequence of the alpha and beta subunits of B-phycoerythrin from the rhodophytan alga Porphyridium cruentum

Determination of the complete amino-acid sequence of the subunits of B-phycoerythrin from Porphyridium cruentum has shown that the alpha subunit contains 164 amino-acid residues and the beta subunit contains 177 residues. When the sequences of B- and C-phycoerythrins are aligned with those of other phycobiliproteins, it is obvious that B-phycoerythrin lacks a deletion at beta-21-22 present in C-phycoerythrin. However, relative to C-phycoerythrin from Fremyella diplosiphon (Calothrix) (Sidler, W., Kumpf, B., Rüdiger, W. and Zuber, H. (1986) Biol. Chem. Hoppe-Seyler 367, 627-642), B-phycoerythrin has deletions at beta-141k-o, beta-142, beta-143, beta-147 and beta-148. The four singly-linked phycoerythrobilins at positions alpha-84, alpha-143a, beta-84 and beta-155, and the doubly-linked phycoerythrobilin at position beta-50/61 are at sites homologous to the attachment sites in C-phycoerythrin. The aspartyl residues (alpha-87, beta-87, and beta-39), that interact with the bilins at alpha-84, beta-84, and beta-155 in C-phycocyanin, are found in the homologous positions in B-phycoerythrin. B-Phycoerythrin, in common with other phycobiliproteins, contains a N gamma-methylasparagine residue at position beta-72.     

In vitro transport of ornithine carbamoyltransferase precursor into rat liver mitochondria using a more homologous medium

The precursor of ornithine carbamoyltransferase can be transported in vitro into rat liver mitochondria using the postmitochondrial supernatant from rat liver, a more homologous medium than the commonly used rabbit reticulocyte lysate. The transport of the precursor in the case of reticulocyte lysate requires a standard translation mixture. In the presence of the postmitochondrial supernatant the same is true. However, when the components of the translation mixture were added individually to the postmitochondrial supernatant, it was found that spermidine or spermine, at physiological concentrations, sufficed for the transport of the precursor of ornithine carbamoyltransferase. The activity of the postmitochondrial supernatant was inactivated by trypsin and slightly decreased by RNase treatment; it was not lost by dialysis or by heating at 100 degrees C.     

Further purification and characterization of the succinyl-CoA:3-hydroxy-3-methylglutarate coenzyme A transferase from rat-liver mitochondria

Succinyl-CoA:3-hydroxy-3-methylglutarate coenzyme A transferase, previously identified in rat-liver mitochondria (Deana et al. (1981), Biochim. Biophys. Acta 662, 119-124), was purified to near homogeneity and further characterized. After the last purification steps consisting of Ultrogel AcA-44 filtration and agarose-hexane-coenzyme A chromatography, the enzyme was apparently tetrameric with a mass of 48-52 kDa determined by gel filtration on Sephadex G-75, ultracentrifugation through a sucrose gradient and SDS-gel electrophoresis. By means of a HPLC technique developed for measuring the CoA esters we could determine the enzyme activity in both forward and reverse directions and show that the kinetic constants, i.e., Km of reactants and Vmax, are not too different for the two reactions. Double-reciprocal plots of the enzyme velocities versus the concentration of one substrate at different fixed concentrations of the other substrate gave families of straight lines converging below the substrate-abscissa for both forward and backward reactions, indicating a kinetic mechanism of rapid equilibrium random Bi-Bi type. The competitive inhibition of the product succinate with respect to both reactants, 3-hydroxy-3-methylglutarate and succinyl-CoA, as well as the Haldane relationships are consistent with this conclusion. An inhibitory effect on CoA transferase activity by acetate, acetoacetate, acetyl-CoA, acetoacetyl-CoA, coenzyme A, carnitine, ZnCl2 and high concentrations of the monovalent anions ClO4-, F-, I- and Cl- was also found.     

Circular pancreatic trypsin inhibitor. A novel substrate for studies on intracellular proteolysis

Several recent studies indicate that substrates for ubiquitin-dependent proteolysis must possess unblocked alpha-amino termini. To examine further the importance of free amino groups for proteolytic susceptibility we selected pancreatic trypsin inhibitor (PTI) as a test substrate. PTI can be circularized to form cPTI, a molecule that lacks alpha-amino groups in the absence of an endoproteolytic cleavage. We compared the breakdown of radioiodinated PTI and cPTI in rabbit reticulocyte lysate and found that cPTI was not stabilized relative to PTI. In addition, proteolysis of PTI or cPTI was not inhibited upon conversion of their lysine residues to homoarginine. However, neither degradation of PTI nor cPTI required ATP, and ubiquitin conjugation to either molecule was minor relative to known substrates of the ubiquitin pathway. Thus, PTI and cPTI are cleaved by an ATP-independent endoprotease(s) that does not require the substrate to be ubiquitinated. Such an activity was identified in low salt fractions obtained upon DEAE chromatography of reticulocyte lysate. The ubiquitin/ATP-dependent protease and another large multisubunit protease, both of which elute from DEAE-Fractogel at higher salt concentrations, do not degrade PTI or cPTI. Although monomeric PTI was rapidly degraded in reticulocyte lysate, cross-linked PTI molecules were stable both in reticulocyte lysate and following introduction into cultured cells using red blood cell-mediated microinjection. Thus, increased rates of turnover do not necessarily correlate with greater molecular mass of the substrate.     

Purification and characterization of DNA-dependent DNA-polymerase alpha from human placenta

A preparation of human placenta DNA polymerase with specific activity 6000 unit/mg was obtained. The protocol of the enzyme purification includes the crude extract preparation, the subsequent chromatographies on phosphocellulose, red sepharose, DEAE sepharose and hydroxylapatite. The isolated DNA polymerase belongs to alpha-type according to the large molecular mass (greater than 150 kDa), high sensitivity to N-ethylmaleimide, the profound inhibition of DNA polymerization activity by 200 mM KCl and the ability to catalyze DNA synthesis, using the deoxyribonucleic template and ribonucleic primer. The DNA polymerase preparations contain a few forms with Stokes radii 50-60 A and sedimentation coefficients 7.3-9.0 S as found from data of gel-filtration and ultracentrifugation in glycerol density gradient, accordingly. The existence of four various forms of DNA polymerase activity: 150, 170, 220, 480 kDa were revealed by native electrophoresis. The four steps of purification result in DNA polymerase preparation that was shown by electrophoresis to contain 15-20% of protein possessing the polymerase activity. However the preparation obtained seems to be a "chromatographically pure substance", according to following ion-exchange and affinity chromatographies. The other proteins without polymerase activity are suggested to be the components of the replicative complex of human placenta cells.     

Isolation of motoneuron cell bodies from spinal cord stored at -70 degrees C in ethylene glycol

Pretreatment of spinal cord with ethylene glycol permits long-term storage of the tissue at -70 degrees C prior to isolation and biochemical analysis of the cell bodies of spinal motoneurons. The method is useful for storing spinal tissue from laboratory animals, as well as from human post mortem specimens, where aliquots of tissue may then be used for motoneuron isolation over an indefinitely long period. In addition to inhibiting the loss of soluble proteins from the neurons during freezing and thawing, cryoprotection increases the yield and improves the appearance of the isolated cell bodies. The method should aid biochemical studies of many kinds of neuronal subpopulations isolated from small amounts of starting material.     

Pattern of changes in clinico-immunological indicators of patients with drug allergy during treatment with autologous blood

An efficient method for the treatment of drug allergy was proposed and practically implemented. The method consists in autologous venous blood being lyzed with sterile bidistilled water at a ratio of 5:1 and injected subcutaneously and partially intradermally into the reflexogenic zones of the back 2-3 cm from the spinal column. During the first week of treatment, increasing doses were injected (3 to 10 ml), whereas during the second week the doses decreased from 10 to 3 ml. Following treatment the patients felt better and featured enhanced working ability as well as markedly declined susceptibility to common colds. The most informative immunological indicators were chosen to evaluate the efficiency of therapy.