Purification of the receptor for platelet-derived growth factor from porcine uterus

The receptor for platelet-derived growth factor has been purified to homogeneity on a large scale from porcine uterus. The purification procedure utilizes solubilization of uterus membranes by Triton X-100, followed by sequential chromatographies on wheat germ agglutinin-Sepharose, fast protein liquid chromatography Mono-Q, and anti-phosphotyrosine-Sepharose. About 160 micrograms of homogeneous and functionally active 170-kDa receptor could be purified from 5 kg of uterus tissue. The pure receptor responded to platelet-derived growth factor stimulation by autophosphorylation, indicating that the receptor has a kinase domain as an integral part of the molecule. A rabbit antiserum was produced against the pure receptor, which specifically recognizes the intact 170-kDa receptor.     

A soluble form of the interleukin 4 receptor in biological fluids

Murine biological fluids and murine cell culture supernatants were analyzed for the presence of soluble murine interleukin 4 receptor (sIL4R) with the use of two monoclonal antibodies directed against the receptor. Mouse urine, serum, ascitic fluid, and cell culture supernatants contained varying levels of immunoreactive protein. All of the immunoreactive protein possessed interleukin 4 (IL 4) binding activity. Following partial purification of ascitic fluid a protein was isolated that binds IL 4 with high affinity. This data is consistent with the fact that murine biological fluids contain a soluble version of the murine IL 4 receptor that arises via secretion of the soluble receptor and/or via shedding of the extracellular portion of the full-length receptor from the cell surface.     

Dicopper(II) and dinickel(II) complexes of Schiff-base macrocycles derived from 5,5-dimethyl-1,9-diformyldipyrromethane

The synthesis and characterisation of two dicopper(II) and two dinickel(II) macrocyclic complexes, [Cu^II₂L^Pr] (10), [Cu^II₂L^Bu] (11), [Ni^II₂L^Pr] (12) and [Ni^II₂L^Bu] (13), are reported. The two new Schiff-base macrocycles (L^Pr)⁴⁻ and (L^Bu)⁴⁻ are isolated as dimetallic complexes 10-13 by the [2+2] condensation of 5,5-dimethyl-1,9-diformyldipyrromethane (9) and 1,3-diaminopropane or 1,4-diaminobutane, respectively, using Cu²⁺ or Ni²⁺ template ions. Single crystal X-ray structure determinations carried out on 10-13 show that each metal atom is in a square planar N₄ geometry, being bound to two deprotonated pyrrole nitrogen atoms of one dipyrromethane unit and to the two adjacent imine nitrogen atoms. NMR spectra obtained for the two dinickel(II) complexes 12 and 13 show that in CDCl₃ solution they are highly symmetrical and diamagnetic.     

Kurze Mitteilungen

Ohne Zusammenfassung     

Risk Stratification of Patients with Peripheral Arterial Disease and Abdominal Aortic Aneurysm Using Aortic Augmentation Index

BackgroundCentral augmentation index (cAIx) is an indicator for vascular stiffness. Obstructive and aneurysmatic vascular disease can affect pulse wave propagation and reflection, causing changes in central aortic pressures.AimTo assess and compare cAIx in patients with peripheral arterial disease (PAD) and / or abdominal aortic aneurysm (AAA).MethodscAIx was assessed by radial applanation tonometry (Sphygmocor) in a total of 184 patients at a tertiary referral centre. Patients were grouped as having PAD only, AAA only, or both AAA and PAD. Differences in cAIx measurements between the three patient groups were tested by non-parametric tests and stepwise multivariate linear regression analysis to investigate associations with obstructive or aneurysmatic patterns of vascular disease.ResultsIn the study sample of 184 patients, 130 had PAD only, 20 had AAA only, and 34 patients had both AAA and PAD. Mean cAIx (%) was 30.5 ± 8.2 across all patients. It was significantly higher in females (35.2 ± 6.1, n = 55) than males (28.4 ± 8.2, n = 129), and significantly higher in patients over 80 years of age (34.4 ± 6.9, n = 22) than in those under 80 years (30.0 ± 8.2, n = 162). Intergroup comparison revealed a significant difference in cAIx between the three patient groups (AAA: 27.3 ± 9.5; PAD: 31.4 ± 7.8; AAA & PAD: 28.8 ± 8.5). cAIx was significantly lower in patients with AAA, higher in patients with both AAA and PAD, and highest in patients with PAD only (beta = 0.21, p = 0.006).ConclusionNon-invasive assessment of arterial stiffness in high-risk patients indicates that cAIx differs according to the pattern of vascular disease. Measurements revealed significantly higher cAIx values for patients with obstructive peripheral arterial disease than for patients with aneurysmatic disease.     

In vitro structural maturation of an early stage pre-40S particle coupled with U3 snoRNA release and central pseudoknot formation

The transition of the 90S to the pre-40S pre-ribosome is a decisive step in eukaryotic small subunit biogenesis leading to a first pre-40S intermediate (state Dis-C or primordial pre-40S), where the U3 snoRNA keeps the nascent 18S rRNA locally immature. We in vitro reconstitute the ATP-dependent U3 release from this particle, catalyzed by the helicase Dhr1, and follow this process by cryo-EM revealing two successive pre-40S intermediates, Dis-D and Dis-E. The latter has lost not only U3 but all residual 90S factors including the GTPase Bms1. In vitro remodeling likewise induced the formation of the central pseudoknot, a universally conserved tertiary RNA structure that comprises the core of the small subunit decoding center. Thus, we could structurally reveal a key tertiary RNA folding step that is essential to form the active 40S subunit.     

The nucleoplasmic phase of pre-40S formation prior to nuclear export

Biogenesis of the small ribosomal subunit in eukaryotes starts in the nucleolus with the formation of a 90S precursor and ends in the cytoplasm. Here, we elucidate the enigmatic structural transitions of assembly intermediates from human and yeast cells during the nucleoplasmic maturation phase. After dissociation of all 90S factors, the 40S body adopts a close-to-mature conformation, whereas the 3'' major domain, later forming the 40S head, remains entirely immature. A first coordination is facilitated by the assembly factors TSR1 and BUD23–TRMT112, followed by re-positioning of RRP12 that is already recruited early to the 90S for further head rearrangements. Eventually, the uS2 cluster, CK1 (Hrr25 in yeast) and the export factor SLX9 associate with the pre-40S to provide export competence. These exemplary findings reveal the evolutionary conserved mechanism of how yeast and humans assemble the 40S ribosomal subunit, but reveal also a few minor differences.     

The use of chemical profiling for monitoring metabolic changes in artificial soil slurries caused by horizontal gene transfer

This study explores the utility of Fourier transform infra-red spectroscopy (FT-IR) as a metabolomic tool to detect changes in water-extractable chemical profile resulting from horizontal gene transfer (HGT) events in artificial soil slurries. A GFP–Km (Green fluorescent protein–kanamycin) cassette tagged HGT recipient Acinetobacter strain ADPWH67 with the salicylate hydroxylase gene (salA) disrupted was introduced to slurries containing either sterile or non-sterile soil. The subsequent addition of naked salA DNA allowed the specific monitoring of HGT events by enumerating GFP-expressing colonies on minimal media with salicylate as a sole carbon source. DNA sequencing confirmed that salA was restored in these transformants. Gene transformation frequencies of around 10⁻⁶ were achieved in the presence of sterile and non-sterile soils. Aqueous extracts of the soil slurries were then analyzed using FT-IR in order to ascertain whether any shifts in chemical profile could be detected. We found that following HGT events FT-IR chemical profiles were clearly altered when analyzed with multivariate statistics. Furthermore, these changes could be explained by differences in key chemical signatures including salicylate as well as other biomolecules found in soils. The slurry extracts were also subjected to GC-MS which confirmed the results of FT-IR analyses. FT-IR was therefore demonstrated to have utility for the rapid screening of metabolomic changes in soils following effective HGT events. In addition, this approach could potentially link specific metabolite changes with corresponding catabolic genes.Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.