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991.
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994.
Ternary complex structure of human HGPRTase, PRPP, Mg2+, and the inhibitor HPP reveals the involvement of the flexible loop in substrate binding.
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G. K. Balendiran J. A. Molina Y. Xu J. Torres-Martinez R. Stevens P. J. Focia A. E. Eakin J. C. Sacchettini S. P. Craig rd 《Protein science : a publication of the Protein Society》1999,8(5):1023-1031
Site-directed mutagenesis was used to replace Lys68 of the human hypoxanthine phosphoribosyltransferase (HGPRTase) with alanine to exploit this less reactive form of the enzyme to gain additional insights into the structure activity relationship of HGPRTase. Although this substitution resulted in only a minimal (one- to threefold) increase in the Km values for binding pyrophosphate or phosphoribosylpyrophosphate, the catalytic efficiencies (k(cat)/Km) of the forward and reverse reactions were more severely reduced (6- to 30-fold), and the mutant enzyme showed positive cooperativity in binding of alpha-D-5-phosphoribosyl-1-pyrophosphate (PRPP) and nucleotide. The K68A form of the human HGPRTase was cocrystallized with 7-hydroxy [4,3-d] pyrazolo pyrimidine (HPP) and Mg PRPP, and the refined structure reported. The PRPP molecule built into the [(Fo - Fc)phi(calc)] electron density shows atomic interactions between the Mg PRPP and enzyme residues in the pyrophosphate binding domain as well as in a long flexible loop (residues Leu101 to Gly111) that closes over the active site. Loop closure reveals the functional roles for the conserved SY dipeptide of the loop as well as the molecular basis for one form of gouty arthritis (S103R). In addition, the closed loop conformation provides structural information relevant to the mechanism of catalysis in human HGPRTase. 相似文献
995.
Bone Matrix Proteins: Isolation and Characterization of a Novel Cell-binding Keratan Sulfate Proteoglycan (Osteoadherin) from Bovine Bone 总被引:6,自引:0,他引:6
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A small cell-binding proteoglycan for which we propose the name osteoadherin was extracted from bovine bone with guanidine hydrochloride–containing EDTA. It was purified to homogeneity using a combination of ion-exchange chromatography, hydroxyapatite chromatography, and gel filtration. The Mr of the proteoglycan was 85,000 as determined by SDS-PAGE. The protein is rich in aspartic acid, glutamic acid, and leucine. Two internal octapeptides from the proteoglycan contained the sequences Glu-Ile-Asn-Leu-Ser-His-Asn-Lys and Arg-Asp-Leu-Tyr-Phe-Asn-Lys-Ile. These sequences are not previously described, and support the notion that osteoadherin belongs to the family of leucine-rich repeat proteins. A monospecific antiserum was raised in rabbits. An enzyme-linked immunosorbent assay was developed, and showed the osteoadherin content of bone extracts to be 0.4 mg/g of tissue wet weight, whereas none was found in extracts of various other bovine tissues. Metabolic labeling of primary bovine osteoblasts followed by immunoprecipitation showed the cells to synthesize and secrete the proteoglycan. Digesting the immunoprecipitated osteoadherin with N-glycosidase reduced its apparent size to 47 kD, thus showing the presence of several N-linked oligosaccharides. Digestion with keratanase indicated some of the oligosaccharides to be extended to keratan sulfate chains. In immunohistochemical studies of the bovine fetal rib growth plate, osteoadherin was exclusively identified in the primary bone spongiosa. Osteoadherin binds to hydroxyapatite. A potential function of this proteoglycan is to bind cells, since we showed it to be as efficient as fibronectin in promoting osteoblast attachment in vitro. The binding appears to be mediated by the integrin αvβ3, since this was the only integrin isolated by osteoadherin affinity chromatography of surface-iodinated osteoblast extracts. 相似文献
996.
Landscape connectivity is a key issue of nature conservation and distance parameters are essential for the calculation of patch level metrics. For such calculations the so-called Euclidean and the least cost distance are the most widespread models. In the present work we tested both distance models for landscape connectivity, using connectivity metrics in the case of a grassland mosaic, and the ground beetle Pterostichus melas as a focal species. Our goal was to explore the dissimilarity between the two distance models and the consequent divergence from the calculated values of patch relevance in connectivity. We found that the two distance models calculated the distances similarly, but their estimations were more reliable over short distances (circa 500 m), than long distances (circa 3000 m). The variability in the importance of habitat patches (i.e. patch connectivity indices) was estimated by the difference between the two distance models (Euclidean vs. least cost) according to the patch size. The location of the habitat patches in the matrix seemed to be a more important factor than the habitat size in the estimation of connectivity. The uncertainty of three patch connectivity indices (Integral Index of Connectivity, Probability of Connectance and Flux) became high above a habitat size of 5 ha. Relevance of patches in maintaining connectivity varied even within small ranges depending on the estimator of distance, revealing the careful consideration of these methods in conservation planning. 相似文献
997.
Odd Nygård Peter Westermann 《Biochimica et Biophysica Acta (BBA) - Gene Structure and Expression》1982,697(3):263-269
An efficient four-step procedure is described for preparing highly purified polypeptide chain initiation factor eIF-3 from rat liver microsomal saltwash. The method involves fractionation with ammonium sulfate between 25–40% saturation (0°C) followed by affinity chromatography on rRNA-cellulose, DEAE-cellulose chromatography and sucrose density gradient centrifugation. eIF-3 is eluted from the affinity column at a KCl concentration of 0.18 M. The purification is 10-times and the recovery of activity better than 85%. In the sucrose gradients, eIF-3 sediments as a 15 S particle indicating a total mass of 650 000 Da. The purified eIF-3 is highly active in stimulating globin synthesis in a fractionated translation system. Factor eIF-3 contains eight subunits with molecular weights ranging from 40 000 to 110 000. Seven of the subunits are present in one copy per eIF-3, whereas the factor contains two copies of one subunit. The isoelectric points of the factor subunits range from 5.5 to 7.3 with most of the polypeptides being acidic. 相似文献
998.
Isolation and properties of a human retinol-transporting protein 总被引:3,自引:0,他引:3
999.
Comparative genetic mapping of cellular rel sequences in man, mouse, and the domestic cat. 总被引:1,自引:0,他引:1
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E Brownell C A Kozak J R Fowle rd W S Modi N R Rice S J O'Brien 《American journal of human genetics》1986,39(2):194-202
We used in situ hybridization techniques to assign the human c-rel locus to the centromere-proximal portion of the short arm of chromosome 2 (2cent-2p13). We also determined the chromosomal location of c-rel sequences in the domestic cat and the laboratory mouse by using a human c-rel fragment to screen panels of rodent X cat and hamster X mouse somatic cell hybrid DNAs. The c-rel locus apparently maintains similar syntenic relationships with other known genetic markers in the human and cat, but displays different linkage relationships in the mouse. 相似文献
1000.
An Evaluation of the MM+ Force Field 总被引:1,自引:0,他引:1
Hyperchem′s MM+ force field, based on Allinger′s MM2 is described and evaluated with respect to other MM2 variants, in terms of rotation barriers, conformational energy differences, and conjugation. Its ability to take missing parameters into account is also evaluated with respect to the Dreiding force field. This evaluation also intends to clearly separate the two different force fields MM+(91) and MM+(**) hiding under the MM+ denomination. It is shown that, whereas MM+ proves to be generally robust, caution must be the rule when dealing with conjugated molecules, particularly with heteroaromatics. 相似文献