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21.
R G Webster G M Air D W Metzger P M Colman J N Varghese A T Baker W G Laver 《Journal of virology》1987,61(9):2910-2916
We previously determined, by X-ray crystallography, the three-dimensional structure of a complex between influenza virus N9 neuraminidase (NA) and the Fab fragments of monoclonal antibody NC-41 [P. M. Colman, W. G. Laver, J. N. Varghese, A. T. Baker, P. A. Tulloch, G. M. Air, and R. G. Webster, Nature (London) 326:358-363, 1987]. This antibody binds to an epitope on the upper surface of the NA which is made up of four polypeptide loops over an area of approximately 600 A2 (60 nm2). We now describe properties of NC-41 and other monoclonal antibodies to N9 NA and the properties of variants selected with these antibodies (escape mutants). All except one of the escape mutants had single amino acid sequence changes which affected the binding of NC-41 and which therefore are located within the NC-41 epitope. The other one had a change outside the epitope which did not affect the binding of any of the other antibodies. All the antibodies which selected variants inhibited enzyme activity with fetuin (molecular weight, 50,000) as the substrate, but only five, including NC-41, also inhibited enzyme activity with the small substrate N-acetylneuramin-lactose (molecular weight, 600). These five probably inhibited enzyme activity by distorting the catalytic site of the NA. Isolated, intact N9 NA molecules form rosettes in the absence of detergent, and these possess high levels of hemagglutinin activity (W.G. Laver, P.M. Colman, R.G. Webster, V.S. Hinshaw, and G.M. Air, Virology 137:314-323, 1984). The enzyme activity of N9 NA was inhibited efficiently by 2-deoxy-2,3-dehydro-N-acetylneuraminic acid, whereas hemagglutinin activity was unaffected. The NAs of several variants with sequence changes in the NC-41 epitope lost hemagglutinin activity without any loss of enzyme activity, suggesting that the two activities are associated with separate sites on the N9 NA head. 相似文献
22.
Sequence and translation of the murine coronavirus 5''-end genomic RNA reveals the N-terminal structure of the putative RNA polymerase. 总被引:20,自引:15,他引:5 下载免费PDF全文
A 28-kilodalton protein has been suggested to be the amino-terminal protein cleavage product of the putative coronavirus RNA polymerase (gene A) (M.R. Denison and S. Perlman, Virology 157:565-568, 1987). To elucidate the structure and mechanism of synthesis of this protein, the nucleotide sequence of the 5' 2.0 kilobases of the coronavirus mouse hepatitis virus strain JHM genome was determined. This sequence contains a single, long open reading frame and predicts a highly basic amino-terminal region. Cell-free translation of RNAs transcribed in vitro from DNAs containing gene A sequences in pT7 vectors yielded proteins initiated from the 5'-most optimal initiation codon at position 215 from the 5' end of the genome. The sequence preceding this initiation codon predicts the presence of a stable hairpin loop structure. The presence of an RNA secondary structure at the 5' end of the RNA genome is supported by the observation that gene A sequences were more efficiently translated in vitro when upstream noncoding sequences were removed. By comparing the translation products of virion genomic RNA and in vitro transcribed RNAs, we established that our clones encompassing the 5'-end mouse hepatitis virus genomic RNA encode the 28-kilodalton N-terminal cleavage product of the gene A protein. Possible cleavage sites for this protein are proposed. 相似文献
23.
Dwight D. Baker 《Physiologia plantarum》1987,70(2):245-248
Fifty strains of Frankia were tested for their ability to nodulate six species of actinorhizal plants. Pure cultured strains were used to inoculate seedlings of Alnus glutinosa (L.) Gaertn., Alnus rubra Bong., Casuarina equisetifolia L., Elaeagnus angustifolia L., Hippophaë rhamnoides L. and Myrica cerifera L. in nutrient solution culture. From the results of this study, host inoculation groups among the actinorhizal plants were defined. Although overlap between host inoculation groups appears to be common, the results from this study did not support the view that Frankia strains are promiscuous. All Frankia strains tested in this study could easily be classified into four major host-specificity groups. 相似文献
24.
Transport and Compartmentation of 1-Aminocyclopropane-1-Carboxylic Acid and Its Structural Analog, alpha-Aminoisobutyric Acid, in Tomato Pericarp Slices 下载免费PDF全文
The uptakes of 1-aminocyclopropane-1-carboxylic acid (ACC), the immediate precursor to ethylene, and its structural analog, α-aminoisobutyric acid (αAIB) by tomato pericarp slices were investigated. Both uptakes show a biphasic (saturable-linear) dependence on external concentration of the transported amino acid. At low concentrations, ACC uptake is competitively inhibited by αAIB and vice versa. Both uptakes also are inhibited by other neutral amino acids but not by acidic or basic amino acids. ACC and αAIB uptakes are metabolically dependent and are increased with time of tissue incubation. αAIB efflux patterns from pericarp slices indicated three distinct αAIB compartments having efflux kinetics consistent with those for cell wall, cytoplasm, and vacuole. The bulk of the αAIB taken up by pericarp tissue is sequestered into the vacuole. The ability of pericarp tissue to accumulate αAIB in the vacuole declines with fruit development. 相似文献
25.
Release of a chimeric protein into the medium from Escherichia coli using the C-terminal secretion signal of haemolysin. 总被引:29,自引:6,他引:23 下载免费PDF全文
Recently, we have identified a novel topogenic sequence at the C terminus of Escherichia coli haemolysin (HlyA) which is essential for its efficient secretion into the medium. This discovery has introduced the possibility of using this secretion system for the release of chimeric proteins from E. coli directly into the medium. We have now successfully fused this C-terminal signal to a hybrid protein containing a few residues of beta-galactosidase and the majority of the E. coli outer membrane porin OmpF lacking its own N-terminal signal sequence. We find that this chimeric protein is specifically translocated across the inner and outer membranes and is released into the medium. In addition, we have further localized the HlyA secretion signal to the final 113 amino acids of the C terminus. In fact, a specific secretion signal appears to reside at least in part within the last 27 amino acids of HlyA. 相似文献
26.
Cartilage proteoglycan aggregates. The link protein and proteoglycan amino-terminal globular domains have similar structures 总被引:10,自引:0,他引:10
Cartilage proteoglycan aggregates contain two components (proteoglycan monomer and link protein) which interact with each other and with hyaluronic acid. Data from amino acid sequence analysis are presented that shows that a domain of the proteoglycan, the hyaluronic acid binding region, which interacts with link protein and hyaluronic acid is very similar to link protein in terms of its primary structure. However, the pattern of glycosylation in the hyaluronic acid binding region is different from that found in link protein. After removal of N-linked oligosaccharides, the tryptically prepared hyaluronic acid binding region from rat chondrosarcoma has a mass by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of 43 +/- 2 kDa. The COOH-terminal two-thirds of rat chondrosarcoma link protein, starting at residue 105, has 41.3% identity with a similar region in the hyaluronic acid binding region. We show that, in addition to the hyaluronic acid binding region, proteoglycan contains another region with similarity to the two repeating loop structures in the COOH-terminal two-thirds of link protein. This presumably corresponds to the second globular domain reported in rotary shadowing studies of cartilage proteoglycans. We have deduced the positions of all of the disulfide bonds in the hyaluronic acid binding region and find them to be in the same positions as would be expected from comparison of these sequences with link protein. 相似文献
27.
The relative contributions of polymer annealing and subunit exchange to microtubule dynamics in vitro 总被引:3,自引:1,他引:2 下载免费PDF全文
Microtubules that are free of microtubule-associated protein undergo dynamic changes at steady state, becoming longer but fewer in number with time through a process which was previously assumed to be based entirely on mechanisms of subunit exchange at polymer ends. However, we recently demonstrated that brain and erythrocyte microtubules are capable of joining end-to-end and suggested that polymer annealing may also affect the dynamic behavior of microtubules in vitro (Rothwell, S. W., W. A. Grasser, and D. B. Murphy, 1986, J. Cell Biol. 102:619-627). In the present study, we first show that annealing is a general property of cytoplasmic microtubules and is not a specialized characteristic of erythrocyte microtubules by documenting annealing between tryosinolated and detyrosinolated brain microtubules. We then examine the contributions of polymer annealing and subunit exchange to microtubule dynamics by analyzing the composition and length of individual polymers in a mixture of brain and erythrocyte microtubules by immunoelectron microscopy. In concentrated preparations of short-length microtubules at polymer-mass steady state, annealing was observed to be the principal factor responsible for the increase in polymer length, whereas annealing and subunit exchange contributed about equally to the reduction in microtubule number. 相似文献
28.
Attachment as a factor in the protection of Enterobacter cloacae from chlorination. 总被引:7,自引:4,他引:3 下载免费PDF全文
Enterobacter cloacae attached to drinking water distribution particles was subjected to chlorination. Attachment resulted in the protection of these organisms from disinfection. This effect was found to be dependent upon both the level of chlorine in the system and attachment time. The results obtained in this study indicate that attached organisms may play an important role in coliform outbreaks. 相似文献
29.
Size of the directing moiety at carbon 5 of cytosine and the activity of human DNA(cytosine-5) methyltransferase 总被引:3,自引:0,他引:3
T A Hardy D J Baker E M Newman L C Sowers M F Goodman S S Smith 《Biochemical and biophysical research communications》1987,145(1):146-152
M13 DNAs in which carbon 5 of each deoxycytidine residue in one strand is replaced with a bulky group are very good substrates for human DNA (cytosine-5) methyltransferase. Rate enhancements of up to 35 fold are obtained depending on the size of the moiety at C-5. The enzyme appears optimally suited to sense a methyl group in one strand at this position. Alkaline density gradient analyses of the distribution of methyl groups applied to 5-BrdCyd or 5-IdCyd substituted DNA reveal that these groups serve to direct the enzyme to methylate the unsubstituted strand. 相似文献
30.
Hen oviduct signal peptidase requires only two proteins for proteolysis of fully synthesized secretory precursor proteins in vitro: one with a molecular mass of 19 kilodaltons (kDa) and one which is a glycoprotein whose mass varies from 22 to 24 kDa depending on the extent of glycosylation. Purified signal peptidase has been analyzed both as part of an active catalytic unit and after electroelution of the individual proteins out of a preparative polyacrylamide gel. The multiple forms of the glycoprotein component of signal peptidase bind to concanavalin A and are shown to be derived from the same polypeptide backbone. Removal of their oligosaccharides by digestion with N-glycanase converts these proteins to a single 19.5-kDa polypeptide. The glycoproteins all exhibit very similar profiles following individual digestion with trypsin and separation of the resulting peptides by reverse-phase high-performance liquid chromatography. In addition, sequence analysis of selected peptides from corresponding regions in chromatograms representing each form of the glycoprotein reveals the same amino acid sequences. The 19-kDa signal peptidase protein does not bind concanavalin A, has a distinct tryptic peptide map from that of the glycoprotein, and appears to share no amino acid sequences in common with the glycoprotein. Its copurification on a concanavalin A-Sepharose column indicates that it must interact directly with the glycoprotein subunit. 相似文献