The use of Taka-diastase in a[3H]poly(A) hybridization assay of oligo(U) sequences in RNA

A reliable assay of uridylate sequences longer than 10 is described. The procedure is based on the hybridization of [3H]poly(A) with poly(U) or oligo(U) sequences in high ionic conditions and a subsequent degradation of single stranded polynucleotides with purified Taka-diastase. A 1:2 complex between poly(A) and poly(U) is formed on which on poly(U) strand is digested by Taka-diastase. The procedure is especially suitable for the detection and quantitation of Un (n greater than 10) in RNA preparation.     

The molecular weight of Artemia ribosomes, as determined from their refractive-index increment and light-scattering intensity

Cytoplasmic ribosomes were isolated from the cryptobiotic embryos of the brine shrimp Artemia salina. Measurements of their refractive-index increments and light-scattering intensities give a value for their molecular weight of (3.4±0.2)×10⁶.     

O-(4-Diazo-3,5-di[125I]iodobenzoyl)sucrose, a novel radioactive label for determining organ sites of catabolism of plasma proteins.

A method is described for radiolabelling proteins with O-(4-diazo-3,5-di[125I]iodobenzoyl)sucrose (DD125IBS). When proteins so labelled were degraded within lysosomes, the radioactive fragments were largely retained within the organelle. High specific radioactivities were obtained without changing the properties of the protein. The validity of the method was demonstrated in vivo in rats using the short-lived protein lactate dehydrogenase, isoenzyme M4, and the long-lived protein bovine serum albumin. Derivatization with DD125IBS did not alter the clearance of either protein. Uptake of DD125IBS-labelled lactate dehydrogenase, isoenzyme M4, by liver and spleen of rats was determined. Radioactivity in these tissues increased up to about 2 h after injection (at this time the protein has been almost completely cleared from the blood) and subsequently declined with a half-life of approx. 20 h. After differential fractionation of liver, radioactivity was largely found in the mitochondrial and lysosomal fraction. The results of these studies establish that DD125IBS covalently coupled to plasma proteins should be a useful radioactive tracer for identifying the tissue and cellular sites of catabolism of relatively long-lived circulating proteins.     

Glucose turnover during pregnancy in anaesthetized post-absorptive rats

During pregnancy the decline in blood [glucose] does not result from the increased distribution space of glucose. The absolute rate of glucose turnover increases in late pregnancy in parallel with the rise in the mass of the conceptus. Nevertheless, glucose turnover per kg body wt. is not increased in late pregnancy, since the lower blood [glucose] decreases glucose utilization by maternal tissues.     

Moulting hormone,juvenile hormone and the ultrastructure of the fat body of adult Sarcophaga bullata (Diptera)

Summary In the ovoviviparous fly, Sarcophaga bullata, vitellogenesis is cyclic; a process reflected in ultrastructural changes in the fat body cells and oenocytes. At eclosion the larval fat body has not yet completely disappeared. During vitellogenesis the fat body cells are specialized for intensive protein synthesis showing a very extensive RER and numerous invaginations of the plasma membrane. These features disappear when the eggs descend into the oviducts to complete embryogenesis. The predominant feature of the oenocytes is their very prominent SER. The fat body cells of the males are never as specialized for protein synthesis as those of the females. Feeding of ecdysterone to males for 3 or more days induces a rather extensive subcellular apparatus for protein synthesis, i.e., invaginations of the plasma membrane and an extensive RER. Juvenile hormone is completely ineffective in this respect. Both ecdysterone and juvenile hormone have pronounced but different effects on the oenocytes of males.     

Gas chromatographic measurement of urinary 5-fluoro-2'-deoxyuridine levels after barium salt precipitation and sephadex LH-20 cleanup

A fluorescent photoaffinity label—8-azido-1-N⁶-etheno-adenosine 5′-triphosphate (8-N₃ε ATP)—for ATP-binding proteins has been synthesized. The effectiveness of the label is demonstrated with F₁ATPase from Micrococcus luteus. 8-N₃ε ATP is a substrate for the enzyme in the presence of bivalent cations. Ultraviolet irradiation of F₁ATPase in the presence of the label and Mg²⁺ ions inhibits the enzyme irreversibly. The fluorescent label is bound preferentially to the β subunit of the enzyme. Labeling and inactivation are decreased by protection with ATP or ADP.     

Branchial Cl transport, anion-stimulated ATPase and acid-base balance inAnguilla anguilla adapted to freshwater: Effects of hyperoxia

Summary Freshwater eel gills are notorious for their limited ability to pump chloride. As a result there is a considerable discrepancy between the Na⁺ and Cl^– plasma levels, and plasma HCO₃ ^– and blood pH are relatively high in this species.When eels are kept in tanks aerated with pure oxygen, significant alterations in blood acid-base balance, an increase in plasma pCO₂ and a decrease in blood pH, are observed. In fish studied after 3 weeks hyperoxia, the decrease in blood pH is compensated by an increase in plasma HCO₃ ^–. Such fish exhibit a Cl^– influx 5 times higher than that observed in normoxic fish. This Cl^– influx is readily inhibited by addition of SCN^– to the external medium.An anion-stimulated ATPase activated by HCO₃ ^– and by Cl^– and inhibited by SCN^– was recently described in membrane fractions of the gills ofCarassius auratus, a fish noted for its high Cl^– pumping rate. This enzyme is also found in the gills of the eel. While the maximal rates of enzyme activation by HCO₃ ^– and by Cl^– are similar inCarassius andAnguilla, the affinity of the enzyme for Cl^– is 25 times higher inCarassius. In the microsomal fraction of the hyperoxic eel gills, the maximal anionstimulated ATPase activity remains unchanged but HCO₃ ^– affinity decreases by 50%, while Cl^– affinity increases 5 times. Thus some characteristics of this ATPase seem to be closely related to the Cl^– pump activity exhibited by the gill in fresh water.     

1H NMR studies at 360 Mhz of the methyl groups in native and chemically modified basic pancreatic trypsin inhibitor (BPTI)

In the ¹H NMR spectra obtained at 360 MHz after digital resolution enhancement, the multiplet resonances of the methyl groups in the basic pancreatic trypsin inhibitor (BPTI) were resolved. With suitable double irradiation techniques the individual methyl resonances were assigned to the different types of aliphatic amino acid residues. Furthermore, from pH titration and comparison of the native protein with chemically modified BPTI, the resonance lines of Ala 16 in the active site and Ala 58 at the C-terminus were identified. Potential applications of the resolved methyl resonances as natural NMR probes for studies of the molecular conformations are discussed.