Molecular dynamics of the 5-HT1a receptor and ligands

A 3-D model of the human 5-HT_1a receptor was constructed fromits amino acid sequence by computer graphics techniques, molecularmechanics calculations and molecular dynamics simulations. Themodel has seven -helical membrane spanning segments, which forma central core containing a putative ligand binding site. Electrostaticpotentials 1.4 Å outside the water accessible surfacewere mainly negative on the synaptic side of the receptor modeland at the postulated ligand binding site, and positive in thecytoplasmic domains. The negative electrostatic potentials aroundthe synaptic domains indicate that positively charged ligandsare attracted to the receptor by electrostatic forces. Moleculardynamics simulations of the receptor model with serotonin, ipsapirone,R(–)-methiothepin or S(+)- methiothepin in the centralcore suggested that up to 22 different amino acid residues mayform a ligand binding pocket, and contribute to the specificityof ligand recognition and binding.     

Methods for phosphate addition in heavy salted cod (Gadus morhua L.)

Headed and gutted fresh or frozen and thawed cod (Gadus morhua L.) from the same net catch were hand filleted post rigor before carrying out small-scale salting trials. Fillets were heavy salted using three different methods introducing phosphate during injection, brining, or during pickle salting. For all salting methods, treatment with 0, 4.5, 9 or 18 g/L of the pyro and tri polyphosphate blend Carnal 2110 was carried out. Quality and chemical parameters were analyzed after 5 weeks and 6 ± 1 months of chilled storage of heavy salted fillets.     

Rehydration of salt-cured cod using injection and tumbling technologies

Traditional rehydration of salt-cured and dried salt-cured cod (klipfish) is most often carried out in stagnant water for 24–48 h. This process has however been difficult to run in industrial scale due to problems related both to process flow and product quality. The new three-step rehydration method developed in this work includes injection of tap water followed by tumbling in water and tumbling in a 2% NaCl brine. Using the new rapid method whole fillets and loins are rehydrated within 3 h. Similar to traditional rehydration, a weight gain of 33–34% was obtained. A trained sensory panel found significant differences in cohesiveness, after-taste and sour smell between fillets rehydrated with the traditional and the new method. No differences were found concerning 14 other sensory properties or the storage stability of rehydrated samples.     

Latitudinal distribution of persistent organic pollutants in pelagic and demersal marine fish on the Norwegian Coast

The latitudinal distribution of persistent organic pollutants (POPs: legacy organochlorines [OCs], polybrominated diphenyl ethers [PBDEs,] and hexabromocyclododecane [HBCD]) was examined in livers of two species of marine fish, the pelagic saithe (Pollachius virens,n = 40) and the demersal cod (Gadus morhua,n = 40), along a south-north gradient (59°-70°N) on the Norwegian Coast. Cod had in general two to three times higher concentrations of POPs than saithe, probably because of higher exposure in the benthic food chain. The concentrations of heavy halogenated compounds were higher in the southernmost region than further north. Moreover, the POP pattern showed a gradual shift in the compositions from south to north, especially for OCs in cod: i.e. the relative importance of low-chlorinated polychlorinated biphenyl (PCB) congeners and some OC-pesticides (e.g., hexachlorobenzen [HCB]) in the contaminant burdens increased with latitude. The latitudinal fractionation signal was weaker in saithe, possibly due to its pelagic and nomadic behavior. Hence, this study shows not only a strong latitudinal fractionation in the compositional patterns of POPs in marine fish but also the effects of habitat use and fish behavior.     

Molecular recognition of docosahexaenoic acid by peroxisome proliferator-activated receptors and retinoid-X receptor alpha

The family of peroxisome proliferator-activated receptors (PPARs) is the molecular target of synthetic antidiabetic and hypolipidemic drugs. The side effects of these drugs are limiting their use in patients with high lipid levels. Natural compounds, like Docosahexaenoic acid (DHA) from fish oil, have beneficial effects in the treatment of metabolic diseases, and several DHA derivatives are known to activate PPAR genes. Experimental studies on affinities of DHA and its derivatives for PPARs are not available. In the present study we are therefore using computational docking, molecular dynamics simulation, and several scoring programs to predict affinities and binding modes of DHA for PPARs and retinoid-X receptor alpha, which is the DNA binding partner of PPARs. The calculations indicated that DHA binds to PPARs and the retinoid-X receptor alpha with high affinity, and that different PPARs exhibited different structural effects on the first four carbons atoms of DHA. Our data indicate that the beneficial health effects of DHA may be obtained by high affinity binding to the PPARs.     

Molecular modelling of UH-301 and 5-HTla receptor interactions

A three-dimensional model of the human 5-HT_1a receptor was constructedby molecular modelling, and the molecular and electronic structuresof (R)- and (S)-5-fluoro-8-hydroxy-2-(dipropylamino)tetralin(UH-301) and of (R)- and (S)-8-hydroxy-2-(dipropylamino)tetralin(8-OH-DPAT) were examined by molecular mechanics and quantummechanics calculations and molecular dynamics simulations. Thereceptor model has seven transmembrane helices (TMHs), organizedaccording to a projection map of visual rhodopsin, and includesall loops between helices and the N- and C-terminal parts. Interactionsof UH-301 and 8-OH-DPAT with the 5-HT_1a receptor were examinedby molecular dynamics simulations and energy minimization ofreceptor–ligand complexes. 8-OH-DPAT had lower electrostaticpotentials around the hydroxyl group and stronger hydrogen bondingto the receptor model than had UH-301. The simulations indicatedthat the 5-HT_1a receptor agonists, (R)- and (S)-8-OH-DPAT and(R)-UH-301, interacted with the receptor at a site closer toAsp82 in TMH2 than did (S)-UH-301, which is a 5-HT,_1a receptorantagonist. Simulations of receptor–ligand complexes indicatedthat Asp82, Asp116, Ser199, Thr200 and De385 are essential forbinding of both agonist and antagonist to the receptor.     

Comparative Molecular Dynamics Simulations of Mitogen-Activated Protein Kinase-Activated Protein Kinase 5

The mitogen-activated protein kinase-activated protein kinase MK5 is a substrate of the mitogen-activated protein kinases p38, ERK3 and ERK4. Cell culture and animal studies have demonstrated that MK5 is involved in tumour suppression and promotion, embryogenesis, anxiety, cell motility and cell cycle regulation. In the present study, homology models of MK5 were used for molecular dynamics (MD) simulations of: (1) MK5 alone; (2) MK5 in complex with an inhibitor; and (3) MK5 in complex with the interaction partner p38α. The calculations showed that the inhibitor occupied the active site and disrupted the intramolecular network of amino acids. However, intramolecular interactions consistent with an inactive protein kinase fold were not formed. MD with p38α showed that not only the p38 docking region, but also amino acids in the activation segment, αH helix, P-loop, regulatory phosphorylation region and the C-terminal of MK5 may be involved in forming a very stable MK5-p38α complex, and that p38α binding decreases the residual fluctuation of the MK5 model. Electrostatic Potential Surface (EPS) calculations of MK5 and p38α showed that electrostatic interactions are important for recognition and binding.     

Salmon farms as a source of organohalogenated contaminants in wild fish

Organohalogenated contaminants (OHCs), including organochlorines (OCs; PCB, and OC-pesticides), brominated flame retardants (BFRs; polybrominated diphenyl ethers [PBDE], hexabromocyclododecane [HBCD]) and perfluorooctane sulfonate (PFOS), were measured in livers of Atlantic cod (Gadus morhua) and saithe (Pollachius virens) caught in the vicinity of salmon farms (n = 75) and control sites (n = 80) in three regions (59°-70°N) in Norway. Forty-five percent of the farm-associated (FA) fish (60% of the saithe and 30% of the cod) and none of the control fish had salmon feed (aquaculture food pellets) in their digestive tracts. Concentrations of OCs and BFRs were about 50% higher and dominated more by persistent compounds in Atlantic cod compared to saithe. After controlling for a set of confounding variables (location, sex, size, weight, gonads size, hepatosomatic index, and % lipids in the liver), the concentrations of ∑OC and ∑BDE were 50% higher in FA cod compared to control fish, whereas they were 20% higher in FA saithe than control fish. Hence, salmon farms are a source of lipid-soluble OHCs to wild marine fish, but variation in life-history and habitat use seems to affect the levels of OHCs in the different fish species. In contrast to the lipid-soluble OHCs, control fish had 67% higher PFOS levels than FA fish, which suggests that natural food contains higher loads of this compound than the commercial feed used in salmon farms. Some OHCs are known to act as endocrine disruptors, thus further work is required to determine if OHCs negatively affect reproductive processes of wild fish associated with salmon farms.     

Survival of Listeria innocua and Listeria monocytogenes in muscle of cod (Gadus morhua L.) during salt-curing and growth during chilled storage of rehydrated product

The aim of this work was to study survival of Listeria innocua and Listeria monocytogenes in muscle of cod during salt-curing and growth during chilled storage of the rehydrated product. Fresh cod was inoculated with L. innocua and L. monocytogenes at different levels before salt-curing. After salt-curing and rehydration, the levels were within 1 log₁₀ CFU/g lower than prior to salt-curing in all experiments. During the first 5 days of storage after rehydration, growth of L. innocua was observed in 1 out of 5 experiments at 4 °C, but a 10–100-fold increase were observed in all experiments from day 5 to day 10. The growth started earlier and was more rapid when samples were stored at 7 °C. Growth of L. monocytogenes at 4 °C appeared to start earlier than for L. innocua, but a 10–100-fold increase was observed also for this bacterium. The lag phases in rehydrated products were longer than in experiments with cod muscle juice. The differences could be explained by a different level of salt stress. This work demonstrates that long term exposure to very high salt concentrations does not eliminate Listeria spp., and that Listeria being present in the fish prior to salt-curing can recover and grow in rehydrated salt-cured cod during chilled storage.     

Origin and spoilage potential of the microbiota dominating genus Psychrobacter in sterile rehydrated salt-cured and dried salt-cured cod (Gadus morhua)

Salt-cured and dried salt-cured cod rehydrated using sterile water and equipment have a short shelf life at 4 degrees C due to high bacterial counts. The microbiota develops off-odours which partly can be described as musty, causing sensory rejection within 7-10 days of chilled storage. The microbiota composition was studied in a total of 38 samples obtained from 10 different, both commercial and laboratory produced, salt-cured and dried salt-cured cod products. The dominating bacterium, representing at least 90% of the total viable count in all products studied, was identified as belonging to the genus Psychrobacter; a Gram-negative, oxidase- and catalase-positive, nonpigmented, halotolerant, psychrotolerant, facultative aerobe and nonmotile bacterium. The morphology of the bacterium resembles coccobacilli and the cells occur most often in pairs. The bacterium was able to hydrolyze lipids, but not proteins. It did not produce H(2)S or TMA and the spoilage in rehydrated salt-cured and dried salt-cured cod is therefor different from what is observed in fresh cod. However, samples inoculated with Psychrobacter immobilis gave the same musty odour as spoiled control samples but earlier in the storage period and of a stronger intensity. In a field experiment, carried out to investigate the origin of the dominating bacterium, it was found that the microbiota in both sterile rehydrated commercially produced and laboratory (aseptically) produced salt-cured cod was dominated by this same bacterium. The bacterium was also isolated from cod skin mucus immediately after capture. The bacterium survived NaCl concentrations up to 25% (w/v) NaCl, stating its ability to survive during the salt-curing process. The dominating bacterium in rehydrated salt-cured and dried salt-cured cod seems to mainly originate from the fresh fish itself and not from contamination during processing.