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871.
Pigeon liver malic enzyme (EC 1.1.1.40) has a double dimer quaternary structure. The NADP+ analogs, aminopyridine adenine dinucleotide phosphate and nicotinamide-1,N6-ethenoadenosine dinucleotide phosphate, bind to the enzyme anti-cooperatively. In the presence of non-cooperative competing ligand NADP+, the binding parameter Hill coefficients of these analogues changed very little. Binding of L-malate with enzyme-AADP+ complex first enhanced then reduced the nucleotide fluorescence. Two L-malate binding sites, with Kd values of 23-30 and 270-400 microM, respectively. for the tight and weak binding sites were postulated. A hybrid model between the sequential and pre-existing asymmetrical models was proposed for the pigeon liver malic enzyme. 相似文献
872.
D J McGillivery W K Yong G G Riffkin B Adler 《International journal for parasitology》1990,20(1):87-93
The 31,000 mol. wt glycoprotein (GP31) antigen of infective third-stage (L3) Ostertagia circumcincta larvae was shown, by surface labelling experiments and immunofluorescent antibody staining of whole larvae and larval sections, to be distributed internally. When transverse sections of L3 O. circumcincta, taken from the anterior pharyngeal region, were further examined by electron microscopy, after immunogold staining with rabbit anti-GP31 antiserum, the GP31 antigen was found to be specifically located in 'secretory organelles' within the cells of the oesophageal glands. By in vitro culturing L3 O. circumcincta in medium supplemented with 35S-methionine and then analysing the excretory-secretory material released by the larvae, it was found that the GP31 molecule was one of the major components of the excretory-secretory complex. The purified GP31 molecule had no detectable proteolytic activity in protein degradation assays. On examination of Triton X-100 extracts of infective larvae from other nematode parasite species, a predominant antigen similar to GP31 was found in Trichostrongylus colubriformis and Haemonchus contortus, but in Toxocara canis a minor component corresponding in mol. wt to GP31 was also detected. Based on these results the possible role of GP31 as a candidate antigen for a broad spectrum molecular vaccine against gastrointestinal nematode parasites in sheep is discussed. 相似文献
873.
UGA-specific nonsense suppressors from Escherichia coli K-12 were isolated and characterized. One of them (Su+UGA-11) was identified as a mutant of the prfB gene for the peptide releasing factor RF2. It appears that in this strain, while peptide release at sites of UGA mutations is retarded, the UGA stop codon is read through even in the absence of a tRNA suppressor, exhibiting a novel type of passive nonsense suppression. Three suppressors (Su+UGA-12, -16 and -34) were capable of restoring the streptomycin sensitive phenotype in resistant bacteria (strAr). Because of their drug-related phenotype, these are possibly mutations in the components of the ribosomal machinery, particularly those concerned with peptide release at UGA nonsense codons. A tRNA suppressor was also obtained which was derived from the tRNA(Trp) gene. In this strain, a long region between rrnC (84.5 min) and rrnB (89.5 min) was duplicated and one of the duplicated genes of tRNA(Trp) was mutated to the suppressor. The mechanism of UGA-suppression is discussed in terms of translation termination at the nonsense codon in both active and passive fashions. 相似文献
874.
N X Zhao M S Ma Y P Zhang D C Xu 《International journal of systematic bacteriology》1990,40(4):452-455
875.
Full-length sequence of the cDNA for human erythroid beta-spectrin 总被引:22,自引:0,他引:22
J C Winkelmann J G Chang W T Tse A L Scarpa V T Marchesi B G Forget 《The Journal of biological chemistry》1990,265(20):11827-11832
Spectrin is the major molecular consituent of the red cell membrane skeleton. We have isolated overlapping human erythroid beta-spectrin cDNA clones and determined 6773 base pairs of contiguous nucleotide sequence. This includes the entire coding sequence of beta-spectrin. The sequence translates into a 2137 amino acid, 246-kDa peptide. beta-Spectrin is found to consist of three distinct domains. Domain I, at the N terminus, is a 272-amino acid region lacking resemblance to the spectrin repetitive motif. Sequences in this region exhibit striking sequence homology, at both nucleotide and amino acid levels, to the N-terminal "actin-binding" domains of alpha-actinin and dystrophin. Between residues 51 and 270 there is 55% amino acid identity to human dystrophin, with only four single amino acid gaps in alignment. Domain II consists of 17 spectrin repeats. Several sequence variations are observed in typical repeat structure. Homology to alpha-actinin extends beyond domain I into the N-terminal portion of domain II. Domain III, 52 amino acid residues at the C terminus, does not adhere to the spectrin repeat motif. Combining knowledge of spectrin primary structure with previously reported functional studies, it is possible to make several inferences regarding structure/function relationships within the beta-spectrin molecule. 相似文献
876.
U J Behrens X L Ma S Bychenok E Baraona C S Lieber 《Biochemical and biophysical research communications》1990,173(1):111-119
Circulating AC levels as well as antibodies against AC-protein adducts are increased in non-alcoholic liver injury. To identify the adducts, we used rats with CCl4-induced cirrhosis. Liver subcellular fractions were analyzed by immunochemical staining of protein slot blots and of electrophoretically separated proteins, transferred to nitrocellulose, using AC-protein adduct-specific antibodies. One reactive protein of about 200 kD was detected in the liver soluble fraction and in the cytosol of isolated hepatocytes and, to a lesser extent in the liver microsomes of CCl4-treated rats; in control animals, this reactivity was much weaker. The immunopositive AC adduct co-migrated with the beta 1,2 dimer of rat collagen type I; it was sensitive to digestion by a highly purified collagenase and also reacted with anti-rat collagen type I-specific IgG. In addition, comparison of peptides of the CNBr-digested, immunoprecipitated AC adduct with those of rat collagen type I revealed a high degree of similarity. Thus, AC adduct formation occurs in liver injury of non-alcoholic origin, and a target protein appears to be related to collagen type I, most likely the procollagen precursor. 相似文献
877.
H S Cheong J S Chang J M Park S M Byun 《Biochemical and biophysical research communications》1990,173(3):795-800
For the enhancement of antibody binding affinity, a bispecific antibody against two different epitopes in human chorionic gonadotropin hormone, one is in alpha-subunit and the other is in beta-subunit, was prepared by chemical recombination using 5,5'-dithiobis(2-nitrobenzoic acid). The epitopes recognized by antibodies were investigated by competitive radioimmunoassay, two-site sandwich radioimmunoassay and additivity assay and a proper epitope pair was chosen for preparation of the bispecific antibody. This bispecific antibody has dual specificity and as much as 17.2-fold higher affinity than that of monoclonal antibody with higher affinity by dual antigen binding radioimmunoassay and Scatchard plot analysis. 相似文献
878.
Site-directed mutagenesis of glycine-14 and two "critical" cysteinyl residues in Drosophila alcohol dehydrogenase 总被引:2,自引:0,他引:2
Three amino acid residues (glycine-14, cysteine-135, and cysteine-218) previously speculated to be important for the structure and function of Drosophila melanogaster alcohol dehydrogenase have been investigated by using site-directed mutagenesis followed by kinetic analysis and chemical modification. Mutating glycine-14 to valine (G14V) virtually inactivates Drosophila ADH, and substitution of alanine at this position (G14A) causes a 31% decrease in activity. Thermal denaturation and kinetic and inhibition studies further demonstrate that replacing glycine-14 with either alanine or valine leads to structural changes in the NAD binding domain. These results provide direct evidence for the role played by glycine-14 in maintaining the correct conformation in the NAD binding domain. On the other hand, changing of cysteine-135, -218, or both to alanine (C135A, C218A, and C135A/C218A) causes no decrease in the catalytic activity of the enzyme, indicating that neither of the cysteinyl residues is essential for catalysis. C135A and wild-type enzyme are both inactivated by DTNB. In contrast, C218A and C135A/C218A are unaffected by DTNB treatment. DTNB modification of cysteine-218 can be prevented by the substrates NAD and 2-propanol, suggesting that cysteine-218 may be in the vicinity of the active site. Cysteine-135 which is normally insensitive to DTNB becomes accessible in the presence of 2-propanol and/or NAD, suggesting a conformational change induced by binding of these substrates. 相似文献
879.
M E Tierno D Mead A E Asato R S Liu N Sekiya K Yoshihara C W Chang K Nakanishi R Govindjee T G Ebrey 《Biochemistry》1990,29(25):5948-5953
Five vinyl-substituted fluororetinal analogues (8-F, 10-F, 12-F, 14-F, and 13,14-F2) were found to give bacteriorhodopsin analogues with properties similar to those of the parent system. Of these, only 14-fluororetinal was found to give an extra red-shifted BR analogue (lambda max less than or equal to 680 nm) in equilibrium with the normal 587-nm pigment. The 680-nm pigment was enriched upon irradiation. It rearranged to the 587-nm pigment at room temperature (delta E [symbol: see text] = 20.8 kcal/mol). Chromophore extraction experiments revealed the all-trans geometry for the 680-nm pigment. 14-Chlororetinal gave a similarly red-shifted pigment while 14-methylretinal did not. A scheme for dark adaptation of the 14-halogenated bacteriorhodopsins has been proposed in which the new red-shifted pigment was assigned the all-trans, 15-syn geometry. 相似文献
880.
Notexin from Notechis scutatus scutatus snake venom was modified with trinitrobenzenesulfonic acid, and the major trinitrophenylated (TNP) derivative was separated by high-performance liquid chromatography. Modification resulted in the incorporation of only one TNP group on the N-terminal alpha-amino group. The TNP derivative showed a precipitous decrease in enzymatic activity and lethal toxicity, whereas the antigenicity remained unchanged. However, trinitrophenylation did not significantly affect the secondary structure of the toxin molecule as revealed by the CD spectra. The results, that the modification reaction was accelerated by the Ca2+ and that the TNP derivative retains its affinity for Ca2+, indicate that the N-terminal alpha-amino group did not participate in the Ca2(+)-binding. The TNP derivative could be regenerated with hydrazine hydrochloride. The biological activities of the regenerated notexin are almost the same as those of native notexin. These results suggest that the N-terminal alpha-amino group is essential for the phospholipase A2 activity and lethal toxicity of notexin, and that incorporation of the TNP group on the N-terminal alpha-amino group might give rise to a distortion of the active conformation of notexin. 相似文献