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41.
U Pande G N La Mar J T Lecomte F Ascoli M Brunori K M Smith R K Pandey D W Parish V Thanabal 《Biochemistry》1986,25(19):5638-5646
The 1H NMR characteristics of the high-spin metmyoglobin from the mollusc Aplysia limacina have been investigated and compared with those of the myoglobin (Mb) from sperm whale. Aplysia metMb exhibits a normal acid----alkaline transition with pK approximately 7.8. In the acidic form, the heme methyl and meso proton resonances have been assigned by 1H NMR using samples reconstituted with selectively deuterated hemins and in the latter case by 2H NMR as well. On the basis of the methyl peak intensities and shift pattern, heme rotational disorder could be established in Aplysia Mb; approximately 20% of the protein exhibits a reversed heme orientation compared to that found in single crystals. Three meso proton resonances have been detected in the upfield region between -16 and -35 ppm, showing that the chemical shift of such protons can serve as a diagnostic probe for a pentacoordinated active site in hemoproteins, as previously shown to be the case in model compounds. The temperature dependence of the chemical shift of the meso proton signals deviates strongly from the T-1 Curie behavior, reflecting the presence of a thermally accessible Kramers doublet with significant S = 3/2 character. Nuclear Overhauser effect, NOE, measurements on Aplysia metMb have provided the assignment of individual heme alpha-propionate resonances and were used to infer spatial proximity among heme side chains. The hyperfine shift values for assigned resonances, the NOE connectivities, and the NOE magnitudes were combined to reach a qualitative picture of the rotational mobility and the orientation of the vinyl and propionate side chains of Aplysia metMb relative to sperm whale MbH2O.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
42.
J. Gos lvez C. L pez-fern ndez J. De La Torre J. A. Suja J. S. Rufas 《Biotechnic & histochemistry》1986,61(4):227-230
A silver staining technique applied to squash preparations of material previously fixed in 3:1 ethanol: acetic acid produces differential staining of the acrosomal region of spermatids during spermiogenesis in orthopteroid species. The method includes treatment with saline sodium citrate solution for 15 min at 60 C, and staining with 50% aqueous silver nitrate adjusted to pH 2.9 with formic acid. 相似文献
43.
The aim of this investigation was to study crypt fission, a process which may be instrumental in regulating crypt number in the intestine. Young Holtzman rats were killed at various times after parturition and samples of the small intestine and colon were removed and processed. A microdissection technique was used to separate crypts from other structures. Crypts were scored as normal or fissioning. The percentage of crypts in fission (PCF) reached peak values of 25% and 52% in the small bowel and colon, respectively, at 21 days post-parturition. From this time onward, the PCF dropped until the adult value of approximately 7% was reached in each site. During this same period, the number of crypts increased from 1.9 X 10(6) to 3.3 X 10(6) in the small bowel and 2.2 X 10(5) to 6.5 X 10(5) in the colon. Thus an inverse relationship between the percentage of crypts in fission and crypt number was found. Distribution of fissure heights in fissioning crypts did not change as the animal aged. The majority of the fissures were found in the lower 1/4 of the fissioning crypts. This suggests that as soon as the fissure extends beyond the stem cell zone, division into two crypts soon occurs. 相似文献
44.
Myoglobin: cytochrome b5 interactions and the kinetic mechanism of metmyoglobin reductase 总被引:2,自引:0,他引:2
D J Livingston S J McLachlan G N La Mar W D Brown 《The Journal of biological chemistry》1985,260(29):15699-15707
Metmyoglobin (metMb) reduction by metMb reductase from heart muscle requires cytochrome b5 as electron-transfer mediator. The existence of a metMb-ferrous cytochrome b5 complex is demonstrated by mutual perturbation of the proteins' respective electrophoretic titration curves between pH 4 and 7. The same technique shows a preferential binding of cytochrome b5 over metMb by the enzyme. The paramagnetic hyperfine shifts in the cytochrome b5 1H NMR spectrum are perturbed by metMb, indicating the formation of a specific bimolecular complex with a 1:1 stoichiometry and a binding constant estimated to be less than 10 microM. The resonances assigned to the cytochrome b5 heme 6-propionate methylene group exhibit the largest complexation shifts. Computer modeling implicates lysines 47, 50, and 98 of metMb as contact points with cytochrome b5 carboxylate residues 43, 44, 60, and heme 6-propionate. The mechanism of the enzymatic reduction establishes metMb reductase as an NADH-cytochrome b5 oxidoreductase. Cytochrome b5 is reduced at near diffusion-controlled rates by the enzyme with a turnover number of 1000 min-1 X Km for the cytochrome is 0.9 microM versus 100 microM reported for the erythrocyte enzyme. Ferrous cytochrome b5 then reduces metMb nonenzymatically with an apparent rate constant of 4.9 X 10(4) M-1 min-1 X Acetylation of metMb, which does not affect its oxygen affinity or chemical reduction, renders it a poor substrate for enzymatic reduction. This study suggests a function for the three exterior lysine residues conserved in all mammalian myoglobin sequences: they are contact points for complexation with cytochrome b5. 相似文献
45.
Among the apoA-I-containing lipoproteins isolated by selected-affinity immunosorption from human serum and plasma, we have identified a subpopulation which, unlike the bulk of high density lipoproteins, has pre-beta electrophoretic mobility. This pre-beta subpopulation can be observed directly in fresh plasma by immunoelectrophoresis. It contains phospholipid and free and esterified cholesterol, but protein constitutes 90% of its mass. Apolipoprotein A-I is the predominant apolipoprotein in this subpopulation; apolipoprotein A-II and the B lipoproteins are not detected. The protein moiety of this subpopulation exhibits markedly lower helicity than that of high density lipoproteins isolated by ultracentrifugation. 相似文献
46.
G Gravante F Sorbera M Sabatino V La Grutta 《Bollettino della Società italiana di biologia sperimentale》1985,61(2):249-254
In chronic free-moving cats unilateral, low frequency, long duration stimulation of the caudate nucleus causes a significant reduction of food intake. The effects are greatest at the beginning of the stimulation. On cessation of stimulation the animals take enough food to compensate for that lost during the time of stimulation. Results are discussed either considering interrelationships between striatum and lateral hypothalamus, either regarding recent viewpoints distinguishing two striatal compartments: dorsal or "non limbic" striatal region and ventral or "limbic" striatal region. 相似文献
47.
48.
49.
Jef D. Boeke Francois La Croute Gerald R. Fink 《Molecular & general genetics : MGG》1984,197(2):345-346
Summary Mutations at the URA3 locus of Saccharomyces cerevisiae can be obtained by a positive selection. Wild-type strains of yeast (or ura3 mutant strains containing a plasmid-borne URA3
+ gene) are unable to grow on medium containing the pyrimidine analog 5-fluoro-orotic acid, whereas ura3
– mutants grow normally. This selection, based on the loss of orotidine-5-phosphate decarboxylase activity seems applicable to a variety of eucaryotic and procaryotic cells. 相似文献
50.