Crystallization of preliminary electron diffraction study to 3.7 A of DNA helix-destabilizing protein gp32*I.

A two-dimensionally large and thin crystal has been obtained from gp32¹I, a proteolytically digested product of a DNA helix-destabilizing protein coded by gene 32 in bacteriophage T4. High-resolution electron diffraction patterns (~3.7Å) are recorded from both unstained and stained protein crystals embedded in glucose. The crystal is of orthorhombic space group with $\text{a = 62.9}\text{A}\text{?}\text{and}\text{b = 47.3}\text{A}\text{?}$.     

Human granulocyte specific nuclear antigen(s). I: Production of antisera and determination of specificity.

Non-histone protein-DNA complexes isolated from human lung tissue were used to immunize New Zealand rabbits. Quantitative microcomplement fixation and immunocytochemistry revealed a specific chromatin antigen(s) in the human granulocyte nucleus. The study demonstrates the feasibility of employing antisera against chromosomal non-histone protein-DNA complexes for the immunochemical identification of cell types.     

Role of calcium in the initiation of fast axonal transport of protein: Effects of divalent cations

Fast axonal transport of [³H]protein has been examined in bullfrog primary afferent neurons incubated in media supplemented with divalent cations that can act as agonists or antagonists of calcium ions. Incubation in calcium-free medium (CFM) had no effect on the rate of transport, but reduced the amount of transported [³H]protein by 40–60% relative to transport in the contralateral preparation maintained in normal medium. Preparations incubated in CFM supplemented with 1.8 mM SrCl₂ (equimolar to the CaCl₂ concentration in normal medium) carried out transport at control levels. Incubation conditions in which primary afferent somata were exposed to the Sr²⁺-medium while nerve trunks were maintained in CFM also supported normal transport. By contrast, selective exposure of nerve trunks to Sr²⁺-medium, and somata to CFM resulted in a reduced level of transport similar to that observed when the whole preparation was incubated in CFM. The depression of transport resulting from incubation in CFM was shown to be reversible when preparations were transferred from CFM to either Sr²⁺-supplemented CFM or to normal medium. By contrast to the effects of Sr²⁺, Ba²⁺ (up to 18 mM) did not substitute for Ca²⁺ in the transport process. When normal medium was supplemented with calciumantagonist cations, the amount of transport was depressed (Co²⁺ > Mn²⁺ >> Mg²⁺), with no concomitant effect on the rate of transport. Results of studies with Co²⁺, as well as those with Sr²⁺, suggest that a major locus of action of these cations is within the neuronal soma at a step subsequent to protein synthesis, and prior to the onset of protein translocation via the transport system. Thus, it is inferred that these divalent cations affect a calcium-dependent step that occurs during the initiation phase of fast axonal transport.     

Internal viscoelastic loading in cat papillary muscle.

The passive mechanical properties of myocardium were defined by measuring force responses to rapid length ramps applied to unstimulated cat papillary muscles. The immediate force changes following these ramps recovered partially to their initial value, suggesting a series combination of viscous element and spring. Because the stretched muscle can bear force at rest, the viscous element must be in parallel with an additional spring. The instantaneous extension-force curves measured at different lengths were nonlinear, and could be made to superimpose by a simple horizontal shift. This finding suggests that the same spring was being measured at each length, and that this spring was in series with both the viscous element and its parallel spring (Voigt configuration), so that the parallel spring is held nearly rigid by the viscous element during rapid steps. The series spring in the passive muscle could account for most of the series elastic recoil in the active muscle, suggesting that the same spring is in series with both the contractile elements and the viscous element. It is postulated that the viscous element might be coupled to the contractile elements by a compliance, so that the load imposed on the contractile elements by the passive structures is viscoelastic rather than purely viscous. Such a viscoelastic load would give the muscle a length-independent, early diastolic restoring force. The possibility is discussed that the length-independent restoring force would allow some of the energy liberated during active shortening to be stored and released during relaxation.     

Effect of a peri fluoro substituent on the conformation of dihydrodiol derivatives of polycyclic aromatic hydrocarbons

$\text{Trans}$-3,4-, 5,6-, 8,9-, and 10,11-dihydrodiols formed from the metabolism of 7-fluorobenz[a]anthracene by rat liver microsomes were isolated by reversed-phase high performance liquid chromatography. Ultraviolet absorption, mass, and NMR spectral analyses indicated that the 5,6- and 8,9-dihydrodiols were preferentially in quasi-diaxial conformations, whereas the 3,4- and 10,11-dihydrodiols were preferentially in quasi-diequatorial conformations. CPK space-filling models suggest that the quasi-diaxial conformation is primarily the result of electronic repulsion between the fluorine and the $\text{peri}$ hydroxyl oxygen. These findings provide a structural basis in the interpretation of the carcinogenic potencies of some fluorinated polycyclic aromatic hydrocarbons.     

The synthesis of 2,1′-anhydro-,2,1′:3,6-dianhydro-, and 2,1′:3,6: 3′,6′-trianhydro-sucrose

1′-O-Mesyl-6,6′-di-O-tritylsucrose and the corresponding 1′-O-tosyl derivative were prepared from 6,6′-di-O-tritylsucrose by selective sulphonylation. Both sulphonates underwent intramolecular cyclisation reactions, to give 2,1′-anhydrosucrose in high yields rather than the isomeric 1′,4′-anhydride. Sequential benzoylation, detritylation, and mesylation of the 2,1′-anhydride afforded 2,1′-anhydro-6,6′-di-O-mesylsucrose tetrabenzoate which, in the presence of base, gave 2,1′:3,6:3′,6′-trianhydrosucrose that was not identical with the product previously claimed to have this structure. Several derivatives of 2,1′-anhydrosucrose were prepared possessing different functional groups at either the 6,6′- or 4,6′-positions. Dimolar mesitylene-sulphonylation of 3,3′,4′6′-tetra-O-acetylsucrose gave the 6,1′-disulphonate, which, in the presence of alkali, gave 2,1′:3,6-dianhydrosucrose, which was transformed into the 2,1′:3,6:3′,6′-trianhydride by sequential bromination at C-6′ (carbon tetrabromide-triphenylphosphine) and base-catalysed cyclisation. Treatment of 3,3′,4′,6′-tetra-O-benzoylsucrose with sulphuryl chloride furnished the 4,6,1′-trichloro derivative, which, on alkaline hydrolysis, was converted into 2,1′:3,6-dianhydro-4-chloro-4-deoxy-galacto-sucrose.     

Role of sodium and water excretion in the antihypertensive effect of vasopressin in the spontaneously hypertensive rat.

Mean arterial pressure (mmHg (1 mmHg = 133.322 Pa)), sodium excretion rate (mumol.kg-1.min-1), and urine flow (microL.kg-1.min-1) were measured in conscious unrestrained spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY) before, during, and after a 3-h intravenous infusion of arginine vasopressin (20 ng.kg-1.min-1), an equipressor dose of phenylephrine, or an infusion of the vehicle. Cessation of the phenylephrine infusion was associated with a return of arterial pressure to preinfusion control values in both SHR and WKY. Cessation of the vasopressin infusion was also associated with a return of arterial pressure to preinfusion values in WKY. In contrast, in the SHR, arterial pressure fell from a preinfusion control level of 164 +/- 6.2 to 137 +/- 4 mmHg within 1 h of stopping the vasopressin infusion. Five hours after stopping the infusion, pressure was 134 +/- 3 mmHg (29 +/- 5 mmHg below preinfusion levels). Similar to the WKY, cessation of a vasopressin infusion was associated with a return of arterial pressure to preinfusion values in Sprague-Dawley rats. Thus, the failure to observe a hypotensive response in normotensive rats was not a peculiarity of the WKY strain. Sodium excretion rates increased during the infusions of vasopressin to a greater extent in SHR than in WKY. However, the natriuresis induced by phenylephrine was not significantly different from that generated by vasopressin in SHR, and in WKY, the natriuresis was greater for phenylephrine than for vasopressin. Urine output increased to a greater extent during the infusions of phenylephrine in both SHR and WKY than during vasopressin infusion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evidence that plasma membrane electrical potential is required for vesicular stomatitis virus infection of MDCK cells: A study using fluorescence measurements through polycarbonate supports

Summary We used fluorescence microscopy of Madin-Darby Canine Kidney (MDCK) cells grown on polycarbonate filters to study a possible link between plasma membrane electrical potential (_pm) and infectivity of vesicular stomatitis virus (VSV). Complete substitution of K⁺ for extracellular Na⁺blocks VSV infection of MDCK cells as well as baby hamster kidney (BHK) cells. When we independently perfused the apical and basal-lateral surfaces of high resistance monolayers, high K⁺ inhibited VSV infection of MDCK cells only when applied to the basal-lateral side; high K⁺ applied apically had no effect on VSV infection. This morphological specificity correlates with a large decrease in _pm of MDCK cells when high K⁺ buffer is perfused across the basal-lateral surface. Depolarization of the plasma membrane by 130 mm basal K⁺ causes a sustained increase of cytosol pH in MDCK cells from 7.3 to 7.5 as reported by the fluorescent dye BCECF. Depolarization also causes a transient increase of cytosol Ca²⁺ from 70 to 300 nm as reported by the dye Fura-2. Neither increase could explain the block of VSV infectivity by plasma membrane depolarization. One alternative hypothesis is that _pm facilitates membrane translocation of viral macromolecules as previously described for colicins, mitochondrial import proteins, and proteins secreted by Escherichia coli.We thank Kenneth Spring for many helpful discussions concerning fluorescence digitized imaging systems, James Russell for his collaboration in the design of our imaging system, Herbert Chase for suggestions on dye loading into MDCK cells, and Manfred Schubert and George Harmison for providing expertise on VSV.