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T Yagi

Publications and source records attributed to T Yagi.

At least 379 records · Page 21Linked to original sources
PubMed1994

Properties of the iron-sulfur center in the 25-kilodalton subunit of the proton-translocating NADH-quinone oxidoreductase of Paracoccus denitrificans.

The 25-kDa subunit of the proton-translocating NADH-quinone oxidoreductase (NDH-1) of Paracoccus denitrificans has been expressed in Escherichia coli and purified to homogeneity. EPR studies of the reduced recombinant protein indicated that the expressed subunit contains a single [2Fe-2S] cluster (Yano, T., Sled', V. D., Ohnishi, T., and Yagi, T. (1994) Biochemistry 33, 494-499). In this report, the electronic, magnetic, and vibrational properties of the [2Fe-2S]2+,+ center have been investigated by the combination of absorption, circular dichroism, variable-temperature magnetic circular dichroism, electron paramagnetic resonance, and resonance Raman spectroscopies and compared with a range of simple [2Fe-2S]-containing proteins. The results are consistent with coordination by two cysteinyl residues at both the reducible and nonreducible iron sites and reveal a striking similarity between the properties of the [2Fe-2S] cluster in the P. denitrificans NDH-1 25-kDa subunit and those of the subclass of ferredoxin-type [2Fe-2S] centers typified by Clostridium pasteurianum 2Fe ferredoxin. The four cyteines residues involved in cluster ligation in these proteins have been tentatively identified based on sequence homology considerations.

Amino Acid Sequence↗
PubMed1994

Mutations in ras genes in cells cultured from mouse skin tumors induced by ultraviolet irradiation.

Mutations in ras oncogenes were detected in cultured cells of mouse skin tumors induced by near-UV irradiation. DNA extracted from the UV-induced tumor cells was transfected to golden hamster embryo cells, and focus-forming ability was confirmed in 22 of 26 cell strains, 15 of which had the repetitive mouse sequence. Mouse ras genes were detected in 10 of these 22 cell strains. Point mutations in the ras genes were at Ha-ras codon 13 (GGC-->GTC in two strains, GGC-->AGC in one strain), Ki-ras codon 61 (CAA-->GAA in two strains), and N-ras codon 61 (CAA-->CAT in two strains, CAA-->AAA in two strains). In one tumor cell strain no base change was directed. Most mutations occurred at dipyrimidine sites. Pyrimidine dimers or pyrimidine(6-4)pyrimidone photoproducts are the likely cause of the skin cancers. The base change occurred preferentially at G.C base pairs, and transversions predominated.

Animals↗
PubMed1994

Initial events of myelination involve Fyn tyrosine kinase signalling.

Myelin is the lipoprotein multimembrane that functions as an insulator preventing the flow of ion currents across the axonal membrane and facilitating the conduction of nerve impulses. It is synthesized by oligodendrocytes in the central nervous system at about the time of birth in mammals. During the initial stages of myelination, several proteins are phosphorylated on tyrosine. Among these proteins, we identified Fyn tyrosine kinase, one of the non-receptor-type tyrosine kinases of the Src family. Here we report that Fyn tyrosine kinase is activated during the initial stages of myelination and that it is associated with the large myelin-associated glycoprotein (MAG), an adhesion molecule that has been implicated in myelinogenesis. The Fyn-large MAG association requires amino-terminal domains of Fyn that include SH2 and SH3 (Src homology domains 2 and 3). Crosslinking of large MAG with antibody induces a rapid increase in the specific activity of Fyn kinase. These results indicate that Fyn participates in the initial events of myelination as a signalling molecule downstream of large MAG; indeed, we find that fyn-deficient mice exhibit impaired myelination.

Animals↗
PubMed1994

Expression of the 25-kilodalton iron-sulfur subunit of the energy-transducing NADH-ubiquinone oxidoreductase of Paracoccus denitrificans.

The energy-transducing NADH-ubiquinone (Q) oxidoreductase of Paracoccus denitrificans is composed of 14 dissimilar subunits and contains at least four iron-sulfur clusters [Yagi, T. (1993) Biochim. Biophys. Acta 1141, 1-17]. The complete DNA sequence of the gene cluster encoding the energy-transducing NADH-Q oxidoreductase of P. denitrificans has been determined. This paper reports the expression of the 25-kilodalton (kDa) (NQO2) subunit of the P. denitrificans enzyme complex in Escherichia coli and the characterization of the iron-sulfur cluster bound to the expressed subunit. The 25-kDa subunit was expressed in the cytoplasmic phase but not in the membrane fraction of E. coli cells and then purified using an affinity nickel chelation column. The purified subunit contains 1.44 mol of non-heme iron and 1.33 mol of acid-labile sulfide/mol of subunit. EPR analysis of the reduced form of this subunit indicates that the expressed subunit contains a single binuclear [2Fe-2S] cluster. This cluster exhibits a spectrum of rhombic symmetry with g values of gx,y,z = 1.913, 1.942, and 1.996, which is very similar to the spectrum of the [2Fe-2S] cluster in the resolved flavoprotein II subfraction (subunit 24 + 9 kDa) of bovine heart complex I [Ragan, C. I., Galante, Y. M., Hatefi, Y., & Ohnishi, T. (1982) Biochemistry 21, 590-594; Ohnishi, T., Ragan, C. I., & Hatefi, Y. (1985) J. Biol. Chem. 260, 2782-2788]. The assignment of the binuclear iron-sulfur cluster of the 25-kDa subunit to an EPR-visible iron-sulfur cluster in the Paracoccus NADH-Q oxidoreductase in situ is discussed.

Amino Acid Sequence↗
PubMed1994

Nanometer scale vibration in mutant axonemes of Chlamydomonas.

Flagellar axonemes of sea urchin sperm display high frequency (200-400 Hz) vibration with nanometer scale amplitudes in the presence of ATP [Kamimura and Kamiya, 1992: J. Cell Biol. 116:1443-1454]. To investigate how various axonemal components affect the vibration, we examined vibration in wild-type and mutant axonemes of Chlamydomonas. At 1 mM ATP, wild-type axonemes underwent vibration at 100-650 Hz with amplitudes of 4-40 nm. This vibration was similar to, but less regular than, that in sea urchin sperm. Axonemes of the mutants ida1 and ida4 lacking part of the inner arm dynein underwent vibrations indistinguishable from that of wild-type. The mutant oda1 lacking the entire outer arm underwent vibration at about half the wild-type frequency. Unexpectedly, the paralyzed mutants pf18 lacking the central pair and pf14 lacking the radial spokes displayed vibration with significantly higher frequencies and smaller amplitudes than those in the wild-type vibration. These results indicate that the high-frequency vibration is common to many kinds of mutant axonemes that lack various axonemal substructures, but that its manner is sensitive to the presence of outer arm dynein and the central pair/radial spoke system. Simultaneous measurements of amplitude and frequency in wild-type and mutant axonemes suggest that the velocity of microtubule sliding in vibrating axonemes is lower than the velocity of sliding under load-free conditions. The velocity is particularly low in pf18.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Far less frequent mutations in ras genes than in the p53 gene in skin tumors of xeroderma pigmentosum patients.

Mutations in Ha-ras, Ki-ras, and N-ras genes in squamous and basal cell carcinomas in patients with xeroderma pigmentosum (XP) were examined by the polymerase chain reaction followed by single-strand conformation polymorphism analysis and direct base sequencing. No mutation was detected in codons 12, 13, and 61 of the ras genes in XP skin tumors. This was in contrast with previous findings of a high frequency of mutation in the p53 gene in skin tumors in XP patients. A novel mutation in codon 6 of the Ki-ras gene was detected in a squamous cell carcinoma. The mutation was a C-->T transition at a dipyrimidine (5'-CT) sequence and could have been produced by solar ultraviolet light. The mutated ras gene did not have the ability to transform NIH/3T3 cells. In three tumors, multiple base substitutions were detected in exon 1 of the Ki-ras and N-ras genes. These results and our previous work on p53 gene mutations suggest that mutations in ras genes are far less frequent than in the p53 gene in the skin tumors in XP patients and that ras genes are less important in skin tumorigenesis in XP patients than is the p53 gene.

3T3 Cells↗
PubMed1994

Increased fearfulness of Fyn tyrosine kinase deficient mice.

Fyn-deficient mice were produced by inserting the beta-galactosidase gene (lacZ) into the fyn gene locus. The homozygously Fyn kinase-deficient (fynz/fynz) mice exhibited stronger light aversion in the light-dark choice test and higher fear-response scores in the novelty preference and passive avoidance tests than did the heterozygously Fyn-deficient (+/fynz) mice. These results indicate that fynz/fynz mice are hyperresponsive to fear-inducing stimuli.

Animals↗
PubMed1994

Functional sites of cytochrome c and other electron carrier proteins: semi-empirical molecular orbital program PM3 applied to the conformational analysis of Cys-X1-X2-Cys peptides.

The semi-empirical molecular orbital program, MOPAC version 6.0, PM3 was applied to estimate the distance between two sulfur atoms of two Cys side chains in optimized conformers of Cys-X1-X2-Cys sequences, as well as the total energy of that conformer relative to the most stable one. Some Cys-X1-X2-Cys tetrapeptides found in cytochrome c were optimized to conformers whose sulfur-sulfur distances were just suitable for binding to heme, whereas some tetrapeptides not found in cytochrome c were unable to be optimized to heme-binding conformers. Similarly, Cys-X1-X2-Cys tetrapeptides found in [4Fe-4S]ferredoxin were optimized to [4Fe-4S]-binding conformers, etc. The tetrapeptides found in the redox site of thioredoxin were optimized to conformers in which the two sulfur atoms were in van der Waals contact, so that a disulfide bond may be formed during the function. The conclusion has been drawn that the combination of X1 and X2 in a Cys-X1-X2-Cys sequence may be determining for that sequence to be a functional redox site in an electron carrier protein.

Amino Acid Sequence↗
PubMed1994

Similarity in the molecular profile of mutations induced by UV light in shuttle vector plasmids propagated in mouse and human cells.

Shuttle vector plasmids pYZ289 were irradiated with UV and transfected to mouse cells to permit repair of damage, mutation and replication in the cells. The frequency and types of mutations were compared with those of UV-irradiated shuttle vector plasmids pZ189 which were propagated in normal human and xeroderma pigmentosum (XP) patient cells defective in DNA repair. Both shuttle vector plasmids contain a bacterial suppressor tRNA gene supF as a common mutation target. pYZ289 propagated in the mouse cells showed survival and mutation frequency similar to pZ189 propagated in the normal human cells. All single base substitution mutations were induced in dipyrimidine sequences and G:C to A:T transition was most frequently observed (47%) in the mouse cells; however, the frequency was significantly lower than that in the XP cells. The frequency of the base substitution mutations at A:T base pairs was significantly higher in the mouse cells (29%) than in the normal human (12%) and the XP cells (6%). These results show that similar types of mutations are induced by UV in mouse and normal human cells, and that the A:T base pair is relatively more mutable in mouse than in normal human and XP cells.

Animals↗
PubMed1994

A novel nonreceptor tyrosine kinase, Srm: cloning and targeted disruption.

We have isolated a novel nonreceptor tyrosine kinase, Srm, that maps to the distal end of chromosome 2. It has SH2, SH2', and SH3 domains and a tyrosine residue for autophosphorylation in the kinase domain but lacks an N-terminal glycine for myristylation and a C-terminal tyrosine which, when phosphorylated, suppresses kinase activity. These are structural features of the recently identified Tec family of nonreceptor tyrosine kinases. The Srm N-terminal unique domain, however, lacks the structural characteristics of the Tec family kinases, and the sequence similarity is highest to Src in the SH region. The expression of two transcripts is rather ubiquitous and changes according to tissue and developmental stage. Mutant mice were generated by gene targeting in embryonic stem cells but displayed no apparent phenotype as in mutant mice expressing Src family kinases. These results suggest that Srm constitutes a new family of nonreceptor tyrosine kinases that may be redundant in function.

Amino Acid Sequence↗
PubMed1994

Ionic conductances of monkey solitary cone inner segments.

1. The membrane properties of cone inner segments dissociated enzymatically from monkey retina were studied under voltage-clamp conditions using patch pipettes in the whole-cell clamp configuration. 2. A noninactivating, voltage-gated calcium current was evoked at potentials positive to -60 mV and peaked between -30 and -20 mV when barium was substituted for calcium. Cadmium (50 microM) but not nickel (50 microM) blocked the current. 3. A large calcium-activated anion current (IAn) was observed when the membrane potential was set to a level between -60 and 30 mV. The reversal potential of IAn was 0 mV with chloride as the sole anion and about -30 and -40 mV when methanesulfonate and D-aspartate, respectively, replaced intracellular chloride to set the equilibrium potential for chloride at -50 mV. IAn inactivated and oscillated when the membrane potential was maintained at depolarized levels, contrary to calcium-activated anionic currents seen in photoreceptors of other species. 4. A sustained-type potassium current was activated by depolarizations positive to -50 mV. The time course of activation and deactivation were voltage dependent. This potassium current was partially blocked by 20 mM tetraethylammonium ions. 5. A transient potassium current was activated by depolarizations positive to -20 mV. This current was blocked by 4-aminopyridine (2 mM) and inactivated with a time constant of approximately 500 ms. The amplitude in response to voltage steps to 45 mV was decreased by prepulses to voltages more positive than -30 mV. 6. Hyperpolarization negative to -65 mV activated an inward current that was completely blocked by external cesium (10 mM). The reversal potential suggested a conductance mechanism permeable to both sodium and potassium ions. 7. A calcium-activated potassium current, which was found in salamander photoreceptors, was not detected. 8. The presence of these conductances is expected to influence the membrane potential and the time course of the light response in monkey cones.

Animals↗
PubMed1994

Immunological approach to Ménière's disease: vestibular immune injury following immune reaction of the endolymphatic sac.

Following direct antigen challenge to the endolymphatic sac (sac), the relation of caloric test results and spontaneous nystagmus to the histological changes of the sensory epithelium of the vestibular organ and perilymph antibody levels was investigated in the guinea pig. In a secondary keyhole limpet hemocyanin (KLH) challenge to the sac, irritative nystagmus was followed by paralytic nystagmus and a suppression of caloric reaction. These findings correlated well with the degree of degeneration of the sensory epithelium of the vestibular organ and the levels of perilymph antibody. On the other hand, neither phosphate-buffered saline inoculation nor primary KLH challenge to the sac nor a secondary KLH challenge to the intradural space resulted in a disturbance of the vestibular function. These results suggest that the immune response of the sac may possibly be an important key to the pathogenesis of Ménière's disease.

Animals↗
PubMed1994

Electrocardiographic diagnosis of left ventricular hypertrophy improved by considering both QRS voltage and ST-T criteria.

We evaluated the usefulness of a combination of QRS voltage and the pattern of ST-T abnormality in the electrocardiographic diagnosis of left ventricular hypertrophy (LVH) in 100 middle-aged men: 32 normals, 59 with hypertension (HT), and 9 with hypertrophic cardiomyopathy (HCM) with evidence of LVH (RV5 or RV6 > or = 2.6 mV, SV1 + RV5 or SV1 + RV6 > or = 3.5 mV). They The subjects were classified into three groups based on ST-T pattern: normal (group N), early strain (group ES), and strain (group S). Echocardiographic evidence of LVH was present in 52.0% (52/100) of the subjects: 72.8% (43/59) of the patients with HT, all 9 patients with HCM, and none of the 32 normals. Echocardiographic evidence of LVH was present in 31.3% (20/64) of group N, 73.3% (11/15) of group ES, and all 21 subjects of group S. In patients with HT, the incidence of echocardiographic LVH was higher in group S (100%) than in both group ES (78.6%) and group N (60.6%). QRS voltage (RV5, RV6, RV5 + SV1, and RV6 + SV1) was significantly correlated with interventricular spetal thickness (IVST), IVST+LVPWT/2, and LV mass, as determined by echocardiography, in patients with LVH (IVST or left ventricular posterior wall thickness (LVPWT) of > or = 12 mm) (r = 0.55 to r = 0.75, p < 0.05), but not in patients without LVH (IVST and LVPWT < 12 mm). There were significant correlations between QRS voltage indices (RV5, RV6, RV5 + SV1, and RV6 + SV1) and IVST, IVST+LVPWT/2, and LV mass in group S (r = 0.68 to r = 0.86, p < 0.05), but not in group N. Values for IVST and LV mass were significantly greater in group S than in groups ES or N. The combination of QRS criteria and ST-T findings reflected the echocardiographic assessment of LVH, especially in patients with HT.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗