A model for the architecture of the hemocyanin from the arthropod Squilla mantis (Crustacea, Stomatopoda)

Squilla mantis hemocyanin is composed of two hexameric subunits but has electron microscopic profiles different from other bis-hexameric hemocyanins, e.g. Astacus and Homarus. We distinguished three different electron microscopic profiles of S. mantis hemocyanin: two sideviews and a topview. These profiles were studied using computer image alignment and correspondence analysis [Van Heel, M. and Frank, J. (1981) Ultramicroscopy 6, 187 - 194]. With the results of this analysis we were able to build a three-dimensional model for the quaternary structure of this hemocyanin. In this model the two hexamers are stacked in such a way that their hexagonal surfaces overlap to about 60% of their width. In the overlap area four subunits are arranged in two different interhexameric pairs, each forming a bridging area between the two hexamers.     

Habitat suitability of the Wadden Sea for restoration of Zostera marina beds

A conceptual model is proposed, describing potential Zostera marina habitats in the Wadden Sea, based on reported data from laboratory, mesocosm and field studies. Controlling factors in the model are dynamics, degree of desiccation, turbidity, nutrients and salinity. A distinction has been made between a higher and a lower zone of potential habitats, each suitable for different morphotypes of Z. marina. The model relates the decline of Z. marina in the Wadden Sea to increased sediment and water dynamics, turbidity, drainage of sediments (resulting in increased degree of desiccation) and total nutrient loads during the twentieth century. The upper and lower delineation of both the higher and the lower zone of potential Z. marina habitats appear to be determined by one or a combination of several of these factors. Environmental changes in one of these factors will therefore influence the borderlines of the zones. The lower zone of Z. marina will be mainly affected by increased turbidity, sediment dynamics, degree of desiccation during low tide and nutrient load. The higher zone will be affected by increases in water and sediment dynamics, desiccation rates and nutrient loads. Potential Z. marina habitats are located above approx. –0.80 m mean sea level (when turbidity remains at the same level as in the early 1990s) in sheltered, undisturbed locations, and preferably where some freshwater influence is present. At locations with a high, near-marine, salinity, the nutrient load has to be low to allow the growth of Z. marina. The sediment should retain enough water during low tide to keep the plants moist. Our results suggest that the return of Z. marina beds within a reasonable time-scale will require not only suitable habitat conditions, but also revegetation measures, as the changes in the environment resulting from the disappearance of Z. marina may impede its recovery, and the natural import of propagules will be unlikely. Furthermore, the lower zone of Z. marina may require a genotype that is no longer found in the Wadden Sea. Received: 26 April 1999 / Received in revised form: 15 October 1999 / Accepted: 16 October 1999     

The p53 isoforms are differentially modified by Mdm2

The discovery that the single p53 gene encodes several different p53 protein isoforms has initiated a flurry of research into the function and regulation of these novel p53 proteins. Full-length p53 protein level is primarily regulated by the E3-ligase Mdm2, which promotes p53 ubiquitination and degradation. Here, we report that all of the novel p53 isoforms are ubiquitinated and degraded to varying degrees in an Mdm2-dependent and -independent manner, and that high-risk human papillomavirus can degrade some but not all of the novel isoforms, demonstrating that full-length p53 and the p53 isoforms are differentially regulated. In addition, we provide the first evidence that Mdm2 promotes the NEDDylation of p53β. Altogether, our data indicates that Mdm2 can distinguish between the p53 isoforms and modify them differently.     

Sows affect their piglets’ faecal microbiota until fattening but not their Salmonella enterica shedding status

Recent studies have shown that Salmonella shedding status affects sows’ microbiota during gestation and that these modifications are reflected in the faecal microbiota of their piglets at weaning. The aims of this study were: (a) to evaluate the persistence, up to the fattening period, of the previously measured link between the microbiota of piglets and their mothers’ Salmonella shedding status; and (b) measure the impact of the measured microbiota variations on their Salmonella excretion at this stage. To achieve this, 76 piglets born from 19 sows for which the faecal microbiota was previously documented, were selected in a multisite production system. The faecal matter of these swine was sampled after 4 weeks, at the fattening stage. The Salmonella shedding status and faecal microbiota of these animals were described using bacteriological and 16S rRNA gene amplicon sequencing respectively. The piglet digestive microbiota association with the Salmonella shedding status of their sows did not persist after weaning and did not affect the risk of Salmonella excretion during fattening, while the birth mother still affected the microbiota of the swine at fattening. This supports the interest in sows as a target for potentially transferrable microbiota modifications.     

Crystal structure of a novel mid-gut procarboxypeptidase from the cotton pest Helicoverpa armigera.

The cotton bollworm Helicoverpa armigera (Hubner) (Lepidoptera: Noctuidae) is one of the most serious insect pests in Australia, India and China. The larva causes substantial economical losses to legume, fibre, cereal oilseed and vegetable crops. This pest has proven to be difficult to control by conventional means, mainly due to the development of pesticide resistance. We present here the 2.5 A crystal structure from the novel procarboxypeptidase (PCPAHa) found in the gut extracts from H. armigera larvae, the first one reported for an insect. This metalloprotease is synthesized as a zymogen of 46.6 kDa which, upon in vitro activation with Lys-C endoproteinase, yields a pro-segment of 91 residues and an active carboxypeptidase moiety of 318 residues. Both regions show a three-dimensional structure quite similar to the corresponding structures in mammalian digestive carboxypeptidases, the most relevant structural differences being located in the loops between conserved secondary structure elements, including the primary activation site. This activation site contains the motif (Ala)(5)Lys at the C terminus of the helix connecting the pro- and the carboxypeptidase domains. A remarkable feature of PCPAHa is the occurrence of the same (Ala)(6)Lys near the C terminus of the active enzyme. The presence of Ser255 in PCPAHa instead of Ile and Asp found in the pancreatic A and B forms, respectively, enlarges the S1' specificity pocket and influences the substrate preferences of the enzyme. The C-terminal tail of the leech carboxypeptidase inhibitor has been modelled into the PCPAHa active site to explore the substrate preferences and the enzymatic mechanism of this enzyme.     

Solid-phase synthesis of polymyxin B1 and analogues via a safety-catch approach.

As part of a program towards the development of novel antibiotics, a convenient method for solid-phase synthesis of the cyclic cationic peptide polymyxin B1 and analogues thereof is described. The methodology, based on cleavage-by-cyclization using Kenner's safety-catch linker, yields crude products with purities ranging from 37-67%. Antibacterial assays revealed that analogues 23-26, in which the (S)-6-methyloctanoic acid moiety is replaced with shorter acyl chains, exhibit distinct antimicrobial activity. The results suggest that the length of the acyl chain is rather critical for antimicrobial activity. On the other hand, substitution of the hydrophobic ring-segment D-Phe-6/Leu-7 in polymyxin B1 with dipeptide mimics (i.e. analogues 27-33) resulted in almost complete loss of antimicrobial activity.     

On the biosynthesis of biotin in Achromobacter IVSW a reinvestigation

[3-¹⁴C, ³⁵S]-L-cysteine was tested as precursor of biotin in Achromobacter IVSW. No significant incorporation was observed, in contradiction with the data previously reported. On the other hand, Achromobacter IVSW converts [³H, ¹⁴C]-dethiobiotin into biotin. This suggests that biotin is biosynthesized in Achromobacter according to the classical dethiobiotin pathway.