The importance of threonine-301 from cytochromes P-450 (laurate (ω-1)-hydroxylase and testosterone 16α-hydroxylase) in substrate binding as demonstrated by site-directed mutagenesis

AbstractThreonine-301 from rabbit liver cytochromes P-450 (laurate (ω-1)-hydroxylase and testosterone 16α-hydroxylase) has been replaced by histidine via site-directed mutagenesis. In the oxidized state the mutant P-450s exhibited typical low-spin type absorption spectra of P-450 and their reduced CO complexes showed a Soret peak at 450 nm. However, no spectral change was induced on addition of substrates for their wild-type counterparts. The mutant P-450s were also completely devoid of the hydroxylase activity. These findings suggest that threonine-301, which is highly conserved in P-450s and located at the distal heme surface, plays an important role in substrate binding

Messung und Optimierung der Lichtpolarisation des A4-Compton-Rückstreupolarimeters

Das A4-Experiment am Teilchenbeschleuniger MAMI der Johannes Gutenberg-Universität Mainz macht sich die Paritätsverletzung in der elastischen Streuung polarisierter Elektronen an unpolarisierten Nukleonen zunutze, um den Beitrag von strange-Quarks zu den elektromagnetischen Vektorformfaktoren des Nukleons zu bestimmen und so zusammen mit den Ergebnissen ähnlicher Experimente an anderen Beschleunigern den Einfluß des "Quark-Sees" auf die statischen Eigenschaften des Nukleons zu beleuchten. Die Meßdaten, die bei Strahlenergien zwischen 315 MeV und 1.5 GeV genommen wurden, können allerdings nur analysiert werden, wenn eine Absolutbestimmung der Strahlpolarisation vorliegt. Zu diesem Zweck wurde ein laserbasiertes Compton-Rückstreupolarimeter entworfen und aufgebaut, mit dem dieser Polarisationsgrad zerstörungsfrei, d.h. parallel zum Experimentierbetrieb und bei denselben Strahlbedingungen wie das Hauptexperiment gemessen werden kann. Ein wichtiger Parameter ist hierbei die Polarisation des Laserstrahls, deren Kenntnis wiederum unabdingbar für die Analyse der Polarimeter-Meßdaten ist. Die hier vorgelegte Arbeit berichtet nun einerseits über die Weiterentwicklung des optischen Systems und seiner Steuerungs-Infrastruktur, andererseits über den Entwurf, die Konstruktion und die Inbetriebnahme eines neuen Meßgeräts zur Bestimmung der Laserpolarisation mitsamt des hierfür erforderlichen Datennahmesystems und der Methode zur Analyse der Meßdaten. Dieses Gerät erreicht eine statistische Meßgenauigkeit von 0.05% bei einer systematischen Unsicherheit von weniger als 0.2%, wobei Veränderungen des Polarisationgrads zwischen dem Ort der Wechselwirkung mit dem Elektronenstrahl und dem Ort der Messung berücksichtigt werden. Die so gewonnenen Daten wurden mit Resultaten einer parallel angefertigten Dissertation kombiniert, um erstmals die mit dem hier beschriebenen Polarimeter während einer Strahlzeit des A4-Experiments gemessene Strahlpolarisation zu extrahieren. Diese konnte trotz Beschränkung auf eine Teilmenge der Daten mit einer statistischen Unsicherheit von 2.5% und einer systematischen Unsicherheit von 0.4% bestimmt werden, was eine deutliche Verbesserung gegenüber dem bisher mit anderen Methoden erzielten Wert von 4% darstellt.The A4 experiment at the Mainz Microtron accelerator facility of the Johannes Gutenberg University in Mainz, Germany, exploits the parity violation in the elastic scattering of polarized electrons off unpolarized nucleons to determine the strange quark contribution to the electromagnetic vector form factors of the nucleon. The objective is to combine these results with those of similar experiments elsewhere in order to shed light on the influence of the "quark sea" on the static properties of the nucleon. In order to analyze the data which were taken at beam energies between 315 MeV and 1.5 GeV, an absolute determination of the beam polarization is necessary. For this purpose, a laser-based Compton backscattering polarimeter, which allows measurement of this polarization parallel to and at the same beam conditions as the main experiment, has been developed and installed in the beamline. One key parameter of this system is the light polarization which must be known in order to interpret the polarimeter data. This work reports on further developments of the optical system and its control infrastructure as well as the design, construction and operation of a novel device for determining the laser polarization including the data acquisition system and analysis methods. Using this device, the polarization state can be measured with 0.05% statistical uncertainty and less than 0.2% systematic uncertainty, wherein changes to the polarization between the electron-photon interaction region and the position of the measuring device are accounted for. These measurements are combined with results from another dissertation in order to extract for the first time the beam polarization measured for the A4 experiment using the Compton polarimeter described here. Despite a limitation to only a subset of the available data, this polarization was determined with 2.5% statistical uncertainty and 0.4% systematic uncertainty, thereby improving on the previous value of 4% achieved using different methods of polarimetry

The Effects of Grain Size and Precipitate on Strength of Nb-Treated Steels

Relationship between the strength and the change in grain size and precipitation behavior of NbC due to heat treatments was examined with 0.2% carbon steels containing 0.02 to 0.09% niobium. The results are summarized as follows. (1) Though NbC particles precipitated in niobium-treated steels tend to coalesce with the rise of austenitizing temperature, they remain in steel and prevent the austenite grain from coarsening because of its small solubility and low dissolving rate in austenite. (2) The highest strength is obtained when the cooling rate from austenitizing temperature to room temperature is 25℃/min. When niobium-treated steel is continuously cooled after a full solution treatment for a long time at sufficiently high temperatures such as 1250℃, NbC seems to precipitate not only at temperatures lower than A_3 point but also at temperatures of austenite range, and this precipitation from austenite will cause less strengthening than that from ferrite. (3) Strengthening effect by niobium-treatment may be explained on the assumption that both effects of the grain refinement and the precipitation of fine particles of NbC are additive to each other. An increase of strength in niobium-treated steels austenitized at temperatures lower than about 1050℃ is due mainly to refining of ferrite grain and that at higher temperatures is due mainly to fine precipitates of NbC

Successful voriconazole treatment of invasive pulmonary aspergillosis in a patient with acute biphenotypic leukemia

A 23-year old woman with acute biphenotypic leukemia (ABL) complained of chest pain with cough, high fever and hemoptysis during induction chemotherapy, although she had been treated with anti-biotics and micafungin. We made a clinical diagnosis of invasive pulmonary aspergillosis (IPA) based on a consolidation in the right upper lung field on a chest radiograph as well as a high level of serum beta-D-glucan (with no evidence of tuberculosis and candidiasis). We changed her treatment from micafungin to voriconazole. Later, we discovered an air-crescent sign by CT scan that supported the diagnosis of IPA. Following voriconazole treatment, clinical symptoms ceased and abnormal chest shadows improved gradually and concurrently with a recovery of neutrophils. IPA must be considered in immunocompromised patients with pulmonary infiltrates who do not respond to broad-spectrum antibiotics. Serological tests and CT findings can aid in early diagnosis of IPA, which, along with treatment for IPA, will improve clinical outcomes.</p

Overexpression of E2F1 associated with LOH at RB locus and hyperphosphorylation of RB in non-small cell lung carcinoma

金沢大学大学院医学系研究科保健学専攻Purpose E2F1 plays a critical role in cell proliferation, and its function is controlled by the retinoblastoma (RB) protein. We examined the expression of E2F1 and the aberration of RB gene and protein to elucidate what factors contribute to the overexpression of E2F1 in non-small cell lung carcinomas. Methods The expression level of E2F1 in tissues of non-small cell lung carcinomas was measured by means of quantitative reverse transcription-polymerase chain reaction and immunohistochemistry. For RB, we examined loss of heterozygosity (LOH) by PCR-restriction fragment length polymorphism and a variable number of tandem repeats, and protein expression by immunohistochemistry. Results Fifteen cases of carcinoma (46%) showed high transcription levels of E2F1 gene. Immunohistochemically, almost all (14 of 15) cases overexpressing E2F1 mRNA were positive for E2F1 protein. LOH at the RB locus was found in 13 of 30 informative cases. In 13 cases with LOH, ten showed overexpression of E2F1 mRNA and protein. Immunohistochemical positivity for phosphorylated RB protein was also closely correlated with overexpression of E2F1. Conclusions Our results suggest that overexpression of E2F1, induced both by LOH at the RB locus and anomalous phosphorylation of the RB protein, is involved in the development of non-small cell lung carcinoma

Lateral–Medial Dissociation in Orbitofrontal Cortex–Hypothalamus Connectivity

The orbitofrontal cortex (OFC) is involved in cognitive functions, and is also closely related to autonomic functions. The OFC is densely connected with the hypothalamus, a heterogeneous structure controlling autonomic functions that can be divided into two major parts: the lateral and the medial. Resting-state functional connectivity has allowed us to parcellate the cerebral cortex into putative functional areas based on the changes in the spatial pattern of connectivity in the cerebral cortex when a seed point is moved from one voxel to another. In the present high spatial-resolution fMRI study, we investigate the connectivity-based organization of the OFC with reference to the hypothalamus. The OFC was parcellated using resting-state functional connectivity in an individual subject approach, and then the functional connectivity was examined between the parcellated areas in the OFC and the lateral/medial hypothalamus. We found a functional double dissociation in the OFC: the lateral OFC (the lateral orbital gyrus) was more likely connected with the lateral hypothalamus, whereas the medial OFC (the medial orbital and rectal gyri) was more likely connected with the medial hypothalamus. These results demonstrate the fundamental heterogeneity of the OFC, and suggest a potential neural basis of the OFC-hypothalamic functional interaction

Adhesiveness of β5 integrin variant lacking FNK767–769 is similar to that of the prototype containing FNKFNK764–769

金沢大学大学院医学系研究科保健学専攻Little is known about the functions of two different β5 integrins: repeated-FNK (FNKFNK764–769) and single-FNK (FNK764–766) amino acid sequences in the cytoplasmic domain. We examined whether they occurred as germ line mutations or somatic mutations associated with neoplastic transformation, and whether there were functional alterations. Out of six cultured cell lines, only KATO-III cells had the single-FNK β5 sequence. The single-FNK β5 was found in 9 out of 79 patients with colon carcinoma, but no somatic mutations were detected in cancerous tissues. CHO cells were transformed with expression vectors containing single-FNK or repeated-FNK β5 cDNA, which were derived from KATO-III cells. CHO cells transfected with single-FNK and repeated-FNK showed similar adhesiveness to, and proliferative activity on, vitronectin substrates