Modelling and parameter identification for batch fermentations with Streptomyces tendae under phosphate limitation

Summary A simple structured model for the dynamics of phosphate-limited batch fermentations with Streptomyces tendae is presented. The model describes the influence of intracellular phosphate storage upon the growth behav our of the culture. The development of the model takes into account the possible internal regulatory processes of phosphate metabolism. These complex biochemical pathways are summarized with regard to rate-limiting steps to obtain relatively simple model equations. The model parameters are fitted to the experimental data with an identification programme based on the sequential quadratic programming algorithm. Modifications in this algorithm yield a good performance for this application. With respect to the sensitivity of the model parameters, a feedback on the modelling is given. After several loops of modelling and identification, a model was achieved that fits to a set of batch fermentations. Furthermore the simulations show that RNA measurements of some recent fermentations can be interpreted by the simulated internal state variable and that there is evidence for RNA as an intracellular phosphate reserve. Offprint requests to: K.-P. Kuhn     

Gene for integrin-associated protein (IAP, CD47): physical mapping, genomic structure, and expression studies in skeletal muscle.

Integrin-associated protein (IAP) is a widely expressed membrane protein with multiple functions in immunological and neuronal processes. Having physically mapped the IAP gene into a BAC/PAC contig covering approximately 1 Mb on human chromosome 3ql3.1-q13.2, we determined the genomic organization of the gene, established its expression in skeletal muscle, and identified a novel splice variant. Our expression studies demonstrate expression of integrin-associated protein in the t-tubular system and the euchromatin of skeletal muscle cells where its function thus far is not known.     

Biliproteidkomposition und Pigmentverteilung in scheibchenförmigen Phycobilisomen von Rhodella violacea

Ein Modell der Anordnung von Phycoerythrin, Phycocyanin und Allophycocyanin in den scheibchenförmigen Phycobilisomen von Rhodella violacea wird vorgestellt; es steht in Einklang mit der quantitativen und qualitativen Pigmentkomposition, dem intermolekularen Energietransfer sowie der Feinstruktur isolierter Phycobilisomen. Hohe Lichtintensitäten und sechstägiger Nitratmangel führen zu einer Reduktion des gesamten Biliproteidgehalts der Zellen. Darüber hinaus ist eine Veränderung der Pigmentkomposition der isolierten Phycobilisomen nachweisbar. Der Phycocyanin- und der Allophycocyaningehalt sind davon unberührt, während der Phycoerythrinanteil auf wenigstens 78% verringert wird. Der Verlust von zwei bis drei Phycoerythrinaggregaten je Phycobilisom beeinträchtigt weder die Fähigkeit zum Energietransfer noch die Fluoreszenzausbeute bei 665 nm in Suspensionen gleichen Phycocyanin- und Allophycocyaningehalts. Wir danken der Deutschen Forschungsgemeinschaft (SFB 103) für die finanzielle Unterstützung.     

Survivability and long-term stress reactivity levels following repeated exposure to nuclear magnetic resonance imaging procedures in rats

The effect of exposure to the magnetic and radio-frequency fields associated with Nuclear Magnetic Resonance Imaging (MRI) on survivability and long-term stress reactivity levels was examined in male rats. Rats in the experimental condition were exposed to MRI for 22.5 minutes for five successive days (Expt. 1) or 23.3 minutes for twenty-one successive days (Expt. 2). Sham field exposed animals received the identical treatment as the exposed animals except that the magnetic and radio-frequency fields were absent. Control rats were also maintained. Thirteen to twenty-two months after the exposure procedure the remaining rats were sacrificed and their whole body, spleen, heart, thymus, and adrenal weights were recorded. Blood samples were taken for measurements of red and white blood cell counts, hemoglobin content, as well as adrenocorticotrophin and corticosterone levels. The results fail to provide any evidence for changes in survivability and long-term stress reactivity levels in rats exposed to MRI and thus give no grounds to challenge the view that MRI is a safe diagnostic procedure.     

Cellular effects and clinical implications of SLC2A3 copy number variation

SLC2A3 encodes the predominantly neuronal glucose transporter 3 (GLUT3), which facilitates diffusion of glucose across plasma membranes. The human brain depends on a steady glucose supply for ATP generation, which consequently fuels critical biochemical processes, such as axonal transport and neurotransmitter release. Besides its role in the central nervous system, GLUT3 is also expressed in nonneural organs, such as the heart and white blood cells, where it is equally involved in energy metabolism. In cancer cells, GLUT3 overexpression contributes to the Warburg effect by answering the cell's increased glycolytic demands. The SLC2A3 gene locus at chromosome 12p13.31 is unstable and prone to non-allelic homologous recombination events, generating multiple copy number variants (CNVs) of SLC2A3 which account for alterations in SLC2A3 expression. Recent associations of SLC2A3 CNVs with different clinical phenotypes warrant investigation of the potential influence of these structural variants on pathomechanisms of neuropsychiatric, cardiovascular, and immune diseases. In this review, we accumulate and discuss the evidence how SLC2A3 gene dosage may exert diverse protective or detrimental effects depending on the pathological condition. Cellular states which lead to increased energetic demand, such as organ development, proliferation, and cellular degeneration, appear particularly susceptible to alterations in SLC2A3 copy number. We conclude that better understanding of the impact of SLC2A3 variation on disease etiology may potentially provide novel therapeutic approaches specifically targeting this GLUT.     

Physiological changes in adult rats exposed to an ELF rotating magnetic field

In three Experiments (I, II, IV), adult male rats, between 115 and 150 days of age were exposed to either a 0.5–3 or 3–30 gauss ELF (0.5 Hz) rotating magnetic field (RMF), for 5, 10, or 26 days. The rats exposed to the RMF for 10 and 26 days averaged significantly (p< 0,05) greater water consumption than controls. The group exposed to the RMF for 5 days also consumed more water, although the difference was not significant. ELF-RMF-exposed rats also showed a progressive decrease (p <0.02) in relative thyroid weights, but increase in body weight gain (p <0.001) up to 10 days of exposure, and increase in testicle weights (p < 0.05) up to 26 days of exposure. No significant differences were found between groups for circulating blood eosinophil counts or relative adrenal weights, although again the differences did increase with duration of exposure. In a fourth Experiment (III), rats that were 80 days of age at the beginning of 21 days of exposure did not show any significant differences in the above measures from the control group. The changes in behavior and physiology associated with ELF exposure is discussed in terms of its effects upon the thyroid and its probable liquid crystalline properties.

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Zusammenfassung In drei Untersuchungen (I, II, IV) wurden männliche, 115–150 Tage alte Ratten entweder einem 0,5–3 oder einem 3–30 Gauss ELF (0,5 Hz) rotierenden magnetischen Feld (RMF) für 5, 10 oder 26 Tage ausgesetzt. Die Ratten, die dem RMF für 10 und 26 Tage ausgesetzt waren, hatten einen signifikant (p < 0,05) höheren Wasserkonsum als die Kontrollgruppe. Auch die Tiere, die dem RMF für 5 Tage ausgesetzt waren, tranken mehr Wasser (p > 0,05). Die exponierten Tiere zeigten auch eine fortschreitende Abnahme (p < 0,02) des relativen Schilddrüsengewichts, dagegen eine grössere Gewichtszunahme (p < 0,001) bei bis zu 10 Tagen Exponierung und Zunahme des relativen Hodengewichts (p < 0,05) bei bis zu 26 Tagen Exponierung. Die Unterschiede in der Zahl der zirkulierenden Esinophilen und im relativen Nebennierengewicht waren nicht signifikant. In der vierten Untersuchung (III) mit Ratten die bei Versuchsbeginn 80 Tage alt waren fanden sich nach 21 Tagen Exponierung keine Unterschiede beim Vergleich mit der Kontrollgruppe. Die physiologischen Veränderungen durch ELF-Exponierung, werden über eine Wirkung auf der Schilddrüse und ihre möglichen flüssigen und kristallinen Eigenschaften erklärt.

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Resume Lors de trois essais (I, II et IV), on a exposé des rats mâles, adultes, âgés de 115 à 150 jours à un champ magnétique tournant (CMT) d'une très basse fréquence (FTB) — de 0,5 Hz — soit de 0,5 à 3 Gauss, soit de 3 à 30 gauss, et cela durant 5, 10 ou 26 jours. Les rats exposés au CMT durant 10 ou 26 jours ont bu plus d'eau que des rats de contrôle et cela de façon significative (p < 0,05). Le groupe exposé au CMT durant 5 jours seulement a aussi bu davantage d'eau, mais la différence n'est alors pas significative. Les rats exposés à la fois au CMT et au FTB ont aussi montré une diminution significative (p < 0,02) du poids relatif de la glande thyroïde, une augmentation du poids du corps (p< 0,001) jusqu'au 10ème jour d'exposition et une augmentation du poids des testicules pour 26 jours d'exposition. On n'a par contre pas constaté de différences significatives entre les groupes quant au nombre d'oesinophiles circulant dans le sang, ni quant au poids des glandes surrénales, bien que ces différences augmentassent avec la durée d'exposition. Dans un quatrième essai (III), des rats âgés de 80 jours au début d'une période d'exposition de 21 jours n'ont présenté aucune différence quant aux paramètres mentionnés ci-dessus. Les changements du comportement et de la physiologie dûs à une exposition au FTB sont discutés par rapport aux répercussions d'une telle exposition sur la thyroïde et ses propriétés liquides et cristalline.

     

Magnetic fields inhibit opioid-mediated ‘analgesic’ behaviours of the terrestrial snail,Cepaea nemoralis

1. The terrestrial snail, Cepaea nemoralis, when placed on a warmed surface (40 degrees C) displays a thermal avoidance behaviour that entails an elevation of the anterior portion of the fully extended foot. The latency of this nociceptive response was increased by the prototypical mu and specific kappa opiate agonists, morphine and U-50, 488H, respectively, in a manner indicative of anti-nociception and the induction of 'analgesia'. Pretreatment with the prototypical opiate antagonist, naloxone, blocked the morphine- and reduced the U-50, 488H-induced analgesia. Naloxone had no effects on the thermal response latencies of saline treated animals. 2. Exposure to either cold (7 degrees C) or warm (38 degrees C) temperature stress increased the nociceptive thresholds of Cepaea in a manner indicative of the induction of 'stress-induced analgesia'. The warm stress-induced analgesia was opioid mediated, being blocked by naloxone, whereas, the cold stress-induced analgesia was insensitive to naloxone. 3. Exposure for 15-30 min to 0.5 Hz weak rotating magnetic fields (1.5-8.0 G) significantly reduced the analgesic effects of the mu and kappa opiate agonists in a manner similar to that observed with naloxone. The magnetic stimuli also inhibited the endogenous opioid mediated warm stress-induced analgesia and significantly reduced the cold stress-induced analgesia. The magnetic stimuli had no evident effects on the nociceptive responses of saline-treated animals. The dihydropyridine (DHP) and non-DHP calcium channel antagonists diltiazem, verapamil. and nifedipine differentially and significantly reduced, while the DHP calcium channel agonist, BAY K8644, significantly enhanced the inhibitory effects of the magnetic fields on morphine-induced analgesia.     

Structure-based prediction of modifications in glutarylamidase to allow single-step enzymatic production of 7-aminocephalosporanic acid from cephalosporin C.

Glutarylamidase is an important enzyme employed in the commercial production of 7-aminocephalosporanic acid, a starting compound in the synthesis of cephalosporin antibiotics. 7-aminocephalosporanic acid is obtained from cephalosporin C, a natural antibiotic, either chemically or by a two-step enzymatic process utilizing the enzymes D-amino acid oxidase and glutarylamidase. We have investigated possibilities for redesigning glutarylamidase for the production of 7-aminocephalosporanic acid from cephalosporin C in a single enzymatic step. These studies are based on the structures of glutarylamidase, which we have solved with bound phosphate and ethylene glycol to 2.5 A resolution and with bound glycerol to 2.4 A. The phosphate binds near the catalytic serine in a way that mimics the hemiacetal that develops during catalysis, while the glycerol occupies the side-chain binding pocket. Our structures show that the enzyme is not only structurally similar to penicillin G acylase but also employs essentially the same mechanism in which the alpha-amino group of the catalytic serine acts as a base. A subtle difference is the presence of two catalytic dyads, His B23/Glu B455 and His B23/Ser B1, that are not seen in penicillin G acylase. In contrast to classical serine proteases, the central histidine of these dyads interacts indirectly with the O(gamma) through a hydrogen bond relay network involving the alpha-amino group of the serine and a bound water molecule. A plausible model of the enzyme-substrate complex is proposed that leads to the prediction of mutants of glutarylamidase that should enable the enzyme to deacylate cephalosporin C into 7-aminocephalosporanic acid.