In tetrapods, only one gene encoding a somatostatin precursor has been identified so far. The present study reports the characterization of the cDNA clones that encode two distinct somatostatin precursors in the brain ofthe frog Rana ridibunda. The cDNAs were isolated by using degenerate oligonucleotides based on the sequence of the central region of somatostatin to screen a frog brain cDNA library. One of the cDNAs encodes a 115-amino acid protein (prepro-somatostatin-14; PSS1) that exhibits a high degree of structural similarity with the mammalian somatostatin precursor. The other cDNA encodes a 103-amino acid protein

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Novel antibacterials are urgently needed to address the growing problem of bacterial resistance to conventional antibiotics. Two-component systems (TCS) are widely used by bacteria to regulate gene expression in response to various environmental stimuli and physiological stress and have been previously proposed as promising antibacterial targets. TCS consist of a sensor histidine kinase (HK) and an effector response regulator. The HK component contains a highly conserved ATP-binding site that is considered to be a promising target for broad-spectrum antibacterial drugs. Here, we describe the identification of putative HK autophosphorylation inhibitors following two independent experimental approaches: in vitro fragment-based screen via differential scanning fluorimetry and in silico structure-based screening, each followed up by the exploration of analogue compounds as identified by ligand-based similarity searches. Nine of the tested compounds showed antibacterial effect against multi-drug resistant clinical isolates of bacterial pathogens and include three novel scaffolds, which have not been explored so far in other antibacterial compounds. Overall, putative HK autophosphorylation inhibitors were found that together provide a promising starting point for further optimization as antibacterials.

The purpose of this study was to examine whether neutral endopeptidase and angiotensin I-converting enzyme, two membrane-bound metalloenzymes that are widely distributed in the microcirculation, play a role in bradykinin-induced increase in vascular permeability in the hamster cheek pouch. Changes in vascular permeability were quantified by counting the number of leaky sites and by calculating the clearance of fluorescein isothiocyanate (FITC)-dextran (molecular mass, 70,000 d) during suffusion of the cheek pouch with bradykinin. Bradykinin produced a concentration-and time-dependent increase in the number of leaky sites and clearance of FITC-dextran. The selective, active site-directed neutral endopeptidase inhibitors phosphoramidon (1.0 gM) and thiorphan (10.0 jiM) and the selective angiotensin I-converting enzyme inhibitor captopril (10.0 ,uM) each shifted the concentration-response curve to bradykinin significantly to the left. During suffusion with bradykinin (1.0 ,uM) and phosphoramidon, the number of leaky sites increased significantly from 17±2 to 27±4 sites per 0.11 cm2 (mean+SEM, p<0.05), and FITC-dextran clearance increased significantly from 1.0±0.2 to 2.1±0.3 ml/secxlO-6. During suffusion with bradykinin (1.0 gM) and captopril, the number of leaky sites increased significantly from 10±2 to 41±3 sites per 0.11 cm2, and FITC-dextran clearance increased significantly from 0.8±0.3 to 3.2±0.8 ml/secxlO-6. During suffusion with bradykinin (1.0 ,LM) and thiorphan, the number of leaky sites increased significantly from 15±3 to 47+7 sites per 0.11 cm2, and FITC-dextran clearance increased significantly from 0.8+0.2 to 4.7+0.6 ml/secxl106. Suffusion (NEP, EC 3.4.24.11) and angiotensin I-converting enzyme (ACE, EC 3.4.15.1), that hydrolyze bradykinin at the Pro7-Phe8 bond to inactive fragnents 1-7 and 8-9.2-4 The location of NEP and ACE in anatomic proximity to the receptors of bradykinin on postcapillary venular endothelial cells suggests that they may play an important role in modulating the edema-forming effects of the peptide in vivo. [2][3][4][5][6][7] We postulated that NEP and ACE each play an important role in modulating the edema-forming effects of bradykinin in vivo. We reasoned that if endogenous NEP and ACE degrade bradykinin to inactive fragments, then selective pharmacological inhibition of

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