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151.
Disulfide and secondary structures of recombinant human granulocyte colony stimulating factor 总被引:4,自引:0,他引:4
Molecular characteristics and secondary structures of recombinant methionyl human granulocyte colony stimulating factor produced by genetically engineered Escherichia coli are described. Limited radiolabeling of the protein with tritiated iodoacetate and determination of the labeled residue revealed that this recombinant protein contains only one free cysteine at position 17 which is not essential for activity. The free cysteine is inaccessible to modification unless the molecule is unfolded under denaturing conditions. The molecule forms two disulfide bridges which were assigned as Cys(36)-Cys(42) and Cys(64)-Cys(74) based on the results of isolation and characterization of disulfide-containing peptides obtained from a subtilisin digest of the intact protein. CD analyses and secondary structure prediction suggest that the molecule is abundant in alpha-helical structures. 相似文献
152.
T C Madhok C C Chao S G Matta A Hong B M Sharp 《Biochemical and biophysical research communications》1989,165(1):151-157
Polyclonal antibodies were raised against a synthetic decapeptide (designated S3) predicted from a segment of the alpha-3 subunit cDNA (amino acid residues 130-139) encoding the rat brain nicotinic cholinergic receptor. This segment was selected because it may be proximate to the nicotine/acetylcholine-binding site of the receptor (1). By radioligand binding assays and sucrose density gradient centrifugation, these monospecific antibodies were shown to inhibit the binding of [3H]nicotine to both the large molecular weight rat brain receptor (240 kDa) and to an SDS-disaggregated nicotine-binding subunit species (80 kDa), in a dose-dependent manner. The neutralizing effect of the anti-S3 antibodies supports the view that this region of the protein is closely related to the agonist binding site. 相似文献
153.
C S Lai J Joseph C C Shih 《Biochemical and biophysical research communications》1989,160(3):1189-1195
We have synthesized a spin-labeled derivative of ET-18-OCH3, a known antitumor ether-linked phospholipid. The spin-labeled analog was shown to be as potent as ET-18-OCH3 in inhibiting 3H-thymidine uptake of HL60 leukemic cells. Electron spin resonance (ESR) studies showed that the mobility of this ether-linked phospholipid in the membrane is more restricted when compared to its ester-linked counterparts. It is probable that the absence of the bulky carbonyl oxygens allows closer packing of the two alkyl chains in the ether-linked phospholipid, thereby reducing the angular amplitude of the motion of the alkyl chains. These findings may be of importance in elucidating mechanisms by which the antitumor ether-linked phospholipids perturb the structure of cellular membranes. 相似文献
154.
155.
31P NMR provides a convenient monitor of the phosphate ester backbone conformational changes upon binding of the intercalating drugs ethidium, quinacrine, and daunomycin to sonicated poly(A).poly(U) and calf thymus DNA. 31P chemical shifts can also be used to assess differences in the duplex unwinding angles in the presence of the drug. Thus a new 31P signal, 1.8-2.2 ppm downfield from the double-stranded helix signals, is observed in the ethidium ion-poly(A).poly(U) complex. This signal arises from phosphates which are in perturbed environments due to intercalation of the drug. This is in keeping with the hypothesis that the P-O ester torsional angle in phosphates linking the intercalated base pairs is more trans-like. Similar though smaller deshielding of the 31P signals is observed in sonicated poly(A).poly(U)-quinacrine complexes as well as in the daunomycin complexes. The effect of added ethidium ion, quinacrine, and daunomycin on the 31P spectra of sonicated calf thymus DNA is consistent with Wilson and Jones' (1982) earlier study. In these drug-DNA complexes the drug produces a gradual downfield shift in the DNA 31P signal without the appearance of a separate downfield peak. These differences are attributed to differences in the rate of chemical exchange of the drug between free and bound duplex states. The previous correlation of 31P chemical shift with drug duplex unwinding angle (Wilson & Jones, 1982) is confirmed for both the RNA and DNA duplexes. 相似文献
156.
Conformational changes of plasma fibronectin detected upon adsorption to solid substrates: a spin-label study 总被引:2,自引:0,他引:2
Changes in local environment of the free sulfhydryl groups in plasma fibronectin upon adsorption of the protein to polystyrene beads have been examined by electron spin resonance (ESR) spin-label spectroscopy. The two free sulfhydryl groups per subunit of plasma fibronectin were modified chemically with an [15N, 2H]maleimide spin-label. For soluble fibronectin, both free sulfhydryl groups are shown to be in confined environments as evidenced from the labeled protein exhibiting a strongly immobilized ESR spectrum as described previously using [14N, 1H]maleimide spin-labels [Lai, C.-S., & Tooney, N. M. (1984) Arch. Biochem. Biophys. 228, 465-473]. When the labeled protein was adsorbed to the beads, half of the strongly immobilized component was found to convert into a weakly immobilized component, a result indicating that one of the two labeled sites becomes exposed and exhibit a fast tumbling motion. Experiments conducted using various spin-labeled fibronectin fragments suggest that the newly exposed labeled site is located between the DNA-binding and the cell-binding regions of the molecule. The data obtained indicate that, upon adsorption to polystyrene beads, plasma fibronectin undergoes a conformational change through which the buried free sulfhydryl group near the cell-binding region of the molecule is exposed. This observation may have important implications regarding the expression of cell adhesive properties of the fibronectin molecule. 相似文献
157.
Acquired immunity against Angiostrongylus cantonensis was induced by immunizing rats with somatic antigens from fifth-stage larvae and adult worms and live third-stage larvae. Rats immunized twice had significantly fewer worms than rats immunized three times. Fewer worms were recovered from rats immunized with 200 live third-stage larvae than from any other groups. Rats immunized with somatic antigens had higher enzyme-linked immunosorbent assay (ELISA) antibody levels than rats immunized with live larvae. Rats immunized with live third-stage larvae of Angiostrongylus cantonensis were more strongly protected against challenge infections (62-92%) than rats immunized with antigens extracted from fifth-stage larvae (0-30%) and adult worms (11-24%). 相似文献
158.
159.
160.
Nine low copy number genomic DNA clones, a ribosomal sequence, and seven cDNA clones were found to identify polymorphisms in cultivated barley (Hordeum vulgare L.). An F2 population consisting of 100 plants was produced from a cross between a high-yielding two-rowed feed barley cultivar and a genetic marker stock homozygous for nine recessive and one dominant morphological marker genes. Through the use of these 10 well-distributed marker genes, five previously mapped isozyme loci, and two storage-protein loci, the approximate recombinational location for each of 17 restriction fragment length polymorphism loci was estimated. One clone, pMSU21, identified variation that appeared to be the result of a small insertion-deletion event that differentiated two-rowed and six-rowed genotypes. This difference was characterized, and one allele was sequenced. Oligonucleotide primers that flanked the insertion-deletion event were synthesized, and DNA samples from the F2 population were subjected to polymerase chain reaction sequence amplification. The variation identified by this technique was determined to be allelic to the variation identified using pMSU21 in Southern blot analysis. Maps of previously undescribed informative clones are included. 相似文献