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T Mikuni

Publications and source records attributed to T Mikuni.

29 records · Page 2Linked to original sources
PubMed1983

Nuclear magnetic resonance studies on calmodulin: spectral assignments in the calcium-free state.

The 400-MHz proton magnetic resonance spectra of calcium-free scallop testis calmodulin (CaM) and pig brain CaM were observed. Detailed spectral assignments were made by resolution enhancement, spin decoupling, and nuclear Overhauser enhancement (NOE) experiments as well as pH titration. Comparison between spectra of scallop testis CaM and pig brain CaM were also utilized for the assignment. Previous assignments for tyrosine-99, histidine-107, epsilon-trimethyllysine-115, and tyrosine-138 [Seamon, K. B. (1980) Biochemistry 19, 207; Krebs, J., & Carafoli, E. (1982) Eur. J. Biochem. 124, 619] were confirmed. Phenylalanine-99 and threonine-143 of scallop testis CaM were identified. Sixteen methyl resonances from one isoleucine, two valines, nine methionines, and the amino-terminal acetyl group were identified. First-stage assignments were made of resonances arising from seven phenylalanines. The uniquely high field shifted phenylalanine resonance previously reported by Seamon was found to consist of two doublets from the two pairs of delta protons of two phenylalanines. The NOE experiments showed that the two phenylalanines are located closely to each other. The large high-field shifts of these phenylalanines were accounted for the ring-current effects due to their proximity. An isoleucine and a valine of which methyl resonances appear at high fields were found to be situated closely to each other. It was found that two delta protons and two epsilon protons of almost all aromatic residues are magnetically equivalent, suggesting that the local structure of aromatic residues is so flexible as to permit the rapid flipping motion of the ring.

Animals↗
PubMed1983

Nuclear magnetic resonance studies on calmodulin: calcium-induced conformational change.

The 400-MHz 1H nuclear magnetic resonance (NMR) studies were carried out on the Ca2+-induced conformational change of calmodulins (CaM's) isolated from scallop testis and pig brain. The resonances were found to be classified approximately into three groups. The resonances of group I, which are perturbed by the binding of Ca2+ to the high-affinity sites, include those of tyrosine-138, epsilon-trimethyllysine-115, histidine-107, tyrosine-99, etc. The previous assignments for tyrosine- (Tyr) 138 [Seamon, K. B. (1980) Biochemistry 19, 207] were corrected. The resonances of group II, which are affected by the binding of Ca2+ to the low-affinity sites, include those of a phenylalanine (Phe), a high field shifted methyl, and a low field shifted alpha-methine. Group III (related to the binding of Ca2+ to both the high-and low-affinity sites) includes the resonances of a Phe, a high field shifted methyl, and threonine-143. It is concluded that sites III and IV are the high-affinity sites. The off-rate of Ca2+ from the high-affinity sites is slower than 50 s-1 while the off-rate from the low-affinity sites is faster than 600 s-1. In the Ca2+-free state, there exists a hydrophobic region containing three phenylalanine (probably Phe-89, Phe-92, and Phe-141), a valine, and an isoleucine in the vicinity of sites III and IV. Tyr-138 is distant from these amino acids. Upon binding of Ca2+ to the high-affinity sites, one of the Phe residues and the valine approach Tyr-138. Similar structural changes were observed between CaM and troponin C when Ca2+ ions are bound to the high-affinity sites. CaM changes in a somewhat different way from troponin C when Ca2+ ions are bound to the low-affinity sites.

Animals↗
PubMedDate not supplied

Hydroxyl free radical production by the reaction of N-methyl-N'-nitro-N-nitrosoguanidine with hydrogen peroxide without exposure to light.

We examined hydroxyl free radical (.OH) production in the mixture of H2O2 and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) without exposure to light using the electron spin resonance spin-trapping technique. When the mixtures were protected from exposure to light, .OH was formed at pH 6.5 and above; it was not formed at pH 5.0 and below, consistent with our previous report. The amount of .OH trapped depended on the concentrations of MNNG and H2O2 and the pH. Nitrite ion was also detected colorimetrically at pH 6.5 and above, but not detected at pH 5.0 and below in the mixtures without exposure to light. Moreover, its production depended on the concentrations of MNNG and H2O2 and the pH. The formation of N-methyl-N'-nitroguanidine in the mixture at pH 7.8 was confirmed by thin-layer chromatography and melting point. These results suggest that nucleophilic attack by H2O2 on the nitroso nitrogen of MNNG results in the formation of N-methyl-N'-nitroguanidine and peroxynitrous acid, which degrades homolytically to yield .OH and nitrogen dioxide, resulting in the production of nitrite ion, at pH 6.5 and above without exposure to light.

Free Radicals↗