Vasopeptidase inhibitors: incorporation of geminal and spirocyclic substituted azepinones in mercaptoacyl dipeptides

A series of 7-(di)alkyl and spirocyclic substituted azepinones were generated and incorporated as conformationally restricted dipeptide surrogates in mercaptoacyl dipeptides. Clear structure-activity relationships with respect to both angiotensin-converting enzyme (ACE) and neutral endopeptidase (NEP) activity in vitro were observed. The best in this series, compound 1g, a geminally dimethylated C-7-substituted azepinone, demonstrated excellent blood pressure lowering in animal models. Compound 1g (BMS-189921) is characterized by a good duration of activity and excellent oral efficacy in models relevant to ACE or NEP inhibition, and its activity is comparable to that of the clinically efficacious agent omapatrilat. Consequently this inhibitor has been advanced clinically for the treatment of hypertension and congestive heart failure.     

Selective Actinide Extraction with a Tri‐Synergistic Mixture Using a Centrifugal Contactor Battery

Abstract

The reprocessing of spent nuclear fuels produces a quantity of high‐level liquid waste (HLLW), which is subsequentely vitrified. The most important contributors to the long‐term radiotoxicity of the vitrified HLLW are the so‐called minor actinides (MAs), which are neptunium, americium, and curium. This paper presents the results obtained during some extraction tests of americium(III) in a synthetic liquid waste representative of that coming from the PUREX and DIAMEX processes, obtained by dissolving the respective nitrate salts of the lanthanides(III) in a diluted solution of nitric acid and spiked with ²⁴¹Am and ¹⁵²Eu. The extraction process employed a tri‐synergistic organic extractant mixture and a centrifugal contactor battery of eight stages.     

Production of surfactant protein C in the baculovirus expression system: the information required for correct folding and palmitoylation of SP-C is contained within the mature sequence

Surfactant protein C (SP-C) is synthesized in the alveolar type II cells of the lung as a 21 kDa propeptide which is proteolytically processed to a 4.2 kDa mature active form. The main function of this extremely hydrophobic protein is to enhance lipid insertion into the air/liquid interface in the lung upon inhalation. This is necessary to maintain a relatively low surface tension at this interface during breathing. In this report we describe the production of mature human SP-C in the baculovirus expression system. The recombinant protein contains a secondary structure with a high alpha-helical content (73%), comparable to native SP-C, as determined by circular dichroism and attenuated total reflection Fourier transform infrared analysis. The expressed protein is a mixture of dipalmitoylated (15%) and non-palmitoylated SP-C. This suggests that the information required for palmitoylation is contained within the sequence of the mature protein. The activity of the protein to insert phospholipids into a preformed monolayer of lipids at an air/liquid interface was determined with a captive bubble surfactometer. Recombinant SP-C significantly reduced the surface tension at the air/liquid interface during dynamic expansion and compression. We conclude that correctly folded, dipalmitoylated and active SP-C can be expressed in the baculovirus expression system. Our results may facilitate investigations into the relation between structure and function of SP-C and into protein palmitoylation in general.     

U1 small nuclear ribonucleoprotein and splicing inhibition by the rous sarcoma virus negative regulator of splicing element

Retroviruses require both spliced and unspliced RNA for replication. Accumulation of unspliced Rous sarcoma virus RNA is facilitated in part by a negative cis element in the gag region, termed the negative regulator of splicing (NRS), which serves to repress splicing of viral RNA but can also block splicing of heterologous introns. The NRS binds components of the splicing machinery including SR proteins, U1 and U2, small nuclear ribonucleoproteins (snRNPs) of the major splicing pathway, and U11 snRNP of the minor pathway, yet splicing does not normally occur from the NRS. A mutation that abolishes U11 binding (RG11) also abrogates NRS splicing inhibition, indicating that U11 is functionally important for NRS activity and suggesting that the NRS is recognized as a minor-class 5' splice site (5' ss). We show here, using specific NRS mutations to disrupt U11 binding and coexpression of U11 snRNA genes harboring compensatory mutations, that the NRS U11 site is functional when paired with a minor-class 3' ss from the human P120 gene. Surprisingly, the expectation that the same NRS mutants would be defective for splicing inhibition proved false; splicing inhibition was as good as, if not better than, that for the wild-type NRS. Comparison of these new mutations with RG11 indicated that the latter may disrupt binding of a factor(s) other than U11. Our data suggest that this factor is U1 snRNP and that a U1 binding site that overlaps the U11 site is also disrupted by RG11. Analysis of mutations which selectively disrupted U1 or U11 binding indicated that splicing inhibition by the NRS correlates most strongly with U1 snRNP. Additionally, we show that U1 binding is facilitated by SR proteins that bind to the 5' half of the NRS, confirming an earlier proposal that this region is involved in recruiting snRNPs to the NRS. These data indicate a functional role for U1 in NRS-mediated splicing inhibition.