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131.
Maternal inheritance of deleted mitochondrial DNA in a family with mitochondrial myopathy 总被引:17,自引:0,他引:17
T Ozawa M Yoneda M Tanaka K Ohno W Sato H Suzuki M Nishikimi M Yamamoto I Nonaka S Horai 《Biochemical and biophysical research communications》1988,154(3):1240-1247
Skeletal muscles from a mother and her daughter both with chronic progressive ophthalmoplegia were analyzed. Histological and biochemical analyses of their muscle samples showed typical features of this type of mitochondrial myopathy. Southern blot analysis revealed that, in both patients, there were two species of mitochondrial DNA (mtDNA): normal one and partially deleted one. The sizes of the deletion were different; the mutant mtDNAs from the mother and the daughter had about 2.5- and 5-kilobase deletions, respectively. The two mutant mtDNAs shared a common deleted region of 1.2-kilobase. However, both the start and the end of deletion were different between them, implying a novel mode of inheritance. This is the first report that the mutant mtDNA is responsible for the maternal inheritance of a human disease. 相似文献
132.
Thermostable alanine racemase from Bacillus stearothermophilus: DNA and protein sequence determination and secondary structure prediction 总被引:5,自引:0,他引:5
The nucleotide sequence of the alanine racemase (EC 5.1.1.1) gene from a thermophile, Bacillus stearothermophilus, was determined by the dideoxy chain termination method with universal and synthetic site-specific primers. The amino acid sequence of the enzyme predicted from the nucleotide sequence was confirmed by peptide sequence information derived from the N-terminal amino acid residues and several tryptic fragments. The alanine racemase gene consists of 1158 base pairs encoding a protein of 386 amino acid residues; the molecular weight of the apoenzyme is estimated as 43,341. The racemase gene of B. stearothermophilus has a closely similar size (1158 vs 1167 base pairs) to that of the gene of a mesophile, B. subtilis, but shows a higher preference for codons ending in G or C. A comparison of the amino acid sequence with those of Bacillus subtilis and Salmonella typhimurium dadB and alr enzymes revealed overall sequence homologies of 31-54%, including an identical octapeptide bearing the pyridoxal 5'-phosphate binding site. Although the residues common in the four racemases are not continuously arrayed, these constitute distinct domains and their hydropathy profiles are very similar. The secondary structure of B. stearothermophilus alanine racemase was predicted from the results obtained by theoretical analysis and circular dichroism measurement. 相似文献
133.
S Nagata K Tanizawa N Esaki Y Sakamoto T Ohshima H Tanaka K Soda 《Biochemistry》1988,27(25):9056-9062
The gene for leucine dehydrogenase (EC 1.4.1.9) from Bacillus stearothermophilus was cloned and expressed in Escherichia coli. The selection for the cloned gene was based upon activity staining of the replica printed E. coli cells. A transformant showing high leucine dehydrogenase activity was found to carry an about 9 kilobase pair plasmid, which contained 4.6 kilobase pairs of B. stearothermophilus DNA. The nucleotide sequence including the 1287 base pair coding region of the leucine dehydrogenase gene was determined by the dideoxy chain termination method. The translated amino acid sequence was confirmed by automated Edman degradation of several peptide fragments produced from the purified enzyme by trypsin digestion. The polypeptide contained 429 amino acid residues corresponding to the subunit (Mr 49,000) of the hexameric enzyme. Comparison of the amino acid sequence of leucine dehydrogenase with those of other pyridine nucleotide dependent oxidoreductases registered in a protein data bank revealed significant sequence similarity, particularly between leucine and glutamate dehydrogenases, in the regions containing the coenzyme binding domain and certain specific residues with catalytic importance. 相似文献
134.
The binding of 25-hydroxy-[26,27-3H]vitamin D-3 and 25-hydroxy-[26,27-3H]vitamin D-2 to the vitamin D binding protein in the plasma of both rats and chicks has been studied. In the case of rats, sucrose density gradient analysis, competitive displacement, and Scatchard analysis demonstrate that 25-hydroxyvitamin D-3 and 25-hydroxyvitamin D-2 are bound equally well to the vitamin D binding protein. In contrast, 25-hydroxyvitamin D-2 is poorly bound, while 25-hydroxyvitamin D-3 is tightly bound to the vitamin D binding protein in chick plasma. On the other hand, the chick intestinal receptor binds 1,25-dihydroxyvitamin D-2 and 1,25-dihydroxyvitamin D-3 equally well with a KD of 7.10(-11) M for both compounds. These results strongly suggest that the failure of the plasma transport protein in chicks to bind the vitamin D-2 compounds may be responsible for their relative ineffectiveness in these animals. 相似文献
135.
Molecular cloning and nucleotide sequence of Thermus thermophilus HB8 trpE and trpG 总被引:11,自引:0,他引:11
The trpE gene of Thermus thermophilus HB8 was cloned by complementation of an Escherichia coli tryptophan auxotroph. The E. coli harboring the cloned gene produced the anthranilate synthase I, which was heat-stable and enzymatically active at higher temperature. The nucleotide sequence of the trpE gene and its flanking regions was determined. The trpE gene was preceded by an attenuator-like structure and followed by the trpG gene, with a short gap between them. No other gene essential for tryptophan biosynthesis was observed after the trpG gene. The amino-acid sequences of the T. themophilus anthranilate synthase I and II deduced from the nucleotide sequence were compared with those of other organisms. 相似文献
136.
Calcium alginate gel stabilized with a polyelectrolyte complex (PEC) consisting of potassium poly(vinyl alcohol) sulfate (KPVS) and trimethylammonium glycol chitosan iodide (TGCI) was used for the immobilization of beta-amylase. The immobilization was made by gelling aqueous droplets of enzyme solution including both sodium alginate and KPVS in a CaCl(2) solution containing TGCI. The activity of the enzyme entrapped into the stabilized gel beads was evaluated by studying the batch reaction kinetics of enzyme-catalyzed hydrolysis of maltotetraose. Repeated kinetic measurements, totaling 18, were carried out at fixed time intervals. After each measurement the beads were stirred for 1 day in a freshly prepared 10 mM NaCl solution at 3 degrees C. It was found that the immobilized system remained stable without leading to a serious loss of the activity or to a large leakage of the enzyme from the support. This was explained as being due to a PEC-crosslinked contracted network structure of the stabilized gel matrix. 相似文献
137.
138.
Effect of pore size in substrate and diffusion of enzyme on hydrolysis of cellulosic materials with cellulases 总被引:1,自引:0,他引:1
The effect of cellulase size on hydrolysis was studied by comparing the behavior of crosslinked cellulase (CC) with normal cellulase (FC). The average molecular weight of the CC was at least three times the molecular weight of the FC. The amounts of each enzyme were adjusted so that the degree of solubilization after 2 h was the same. The degree of solubilization of Avicel with CC was higher than that with FC in the late stage of reaction. The degree of solubilization of pretreated lignocelluloses was much greater than that of Avicel, but the degree of solubilization with CC was lower than that with FC at all times during the reaction. The degree of solubilization of artificial lignified Avicel was higher with FC than with CC, but the degree of solubilization of de-lignified the artificial lignified Avicel was lower with FC than with CC. The degree of solubilization of amorphous celloulose with FC was the same as that with CC at all times during the reaction. These behaviors are examined by the hypothesis that when small pores dominate, the smaller enzyme components diffuse into the pores and become inactive since synergism with the larger components is no longer possible, whereas, when larger pores dominate, the entire enzyme can diffuse in and therefore the available surface area is increased. This hypothesis is supported by direct measurement of the pore size in two of the substrates and by diffusion inside Avicel of only smaller molecular cellulase component. 相似文献
139.
Binding of [3H]befunolol to beta-adrenoceptors in cardiac muscles of fetal and neonatal rat 总被引:1,自引:0,他引:1
K Koike H Tanaka K Shigenobu I Takayanagi 《Canadian journal of physiology and pharmacology》1988,66(7):957-960
Characterization of beta-adrenoceptors was studied in heart muscles of rat fetus and neonate. The results of binding assay with [3H]befunolol, a beta-adrenergic partial agonist, to membrane fractions from rat heart muscles indicate that beta-adrenoceptors contain two different affinity sites. In the presence of 5'-guanylylimidodiphosphate, the low affinity site was reduced, while the high affinity site was not affected. The dissociation constants for both sites did not change during pre- and post-natal development. But the maximum binding sites for both sites decreased slightly but significantly (p less than 0.05) during development. A 10-fold decrease in norepinephrine sensitivity and isoprenaline sensitivity during pre- and post-natal development was not explained by the slight decrease in the maximum binding sites. 相似文献
140.
Thermostable alanine racemase from Bacillus stearothermophilus: molecular cloning of the gene, enzyme purification, and characterization 总被引:5,自引:0,他引:5
The alanine racemase (EC 5.1.1.1) gene of a thermophilic bacterium, Bacillus stearothermophilus, was cloned and expressed in Escherichia coli C600 with vector plasmid pICR301, which was constructed from pBR322 and the L-alanine dehydrogenase gene derived from B. stearothermophilus. A coupled assay method with L-alanine dehydrogenase and tetrazolium salts was used to detect visually the alanine racemase activity in the clones. Alanine racemase overproduced in a clone carrying the plasmid pICR4, 12 kilobases of DNA, was purified from cell extracts about 340-fold to homogeneity by five steps including heat treatment. The overproduced enzyme was confirmed to originate from B. stearothermophilus by an immunochemical cross-reaction with the enzyme of B. stearothermophilus. The purified enzyme has a molecular weight of about 78 000 and consists of two identical subunits of Mr of 39 000. At the optimum temperature (50 degrees C), the enzyme has a specific activity of 1800 units/mg (Vmax, D- to L-alanine). Resolution and reconstitution experiments together with the absorption spectrum of the enzyme clearly indicate that alanine racemase of B. stearothermophilus is a pyridoxal 5'-phosphate enzyme. 相似文献