Ten, electrophoretically distinct, molecular forms of alcohol dehydrogenase have been isolated from a single human liver by affinity and ion-exchange chromatography. The starch gel electrophoresis ...patterns after the dissociation-recombination of the forms are consistent with the hypothesis that they arise from the random combination of alpha, beta 1, gamma 1, and gamma 2 subunits into six heterodimeric and four homodimeric isoenzymes. Large differences in kinetic properties are observed for the homodimeric isoenzymes, alpha alpha, beta 1 beta 1, gamma 1 gamma 1, and gamma 2 gamma 2. At pH 7.5, the Km value of beta 1 beta 1 for ethanol is 0.049 mM and that of alpha alpha is 4.2 mM. Forms gamma 1 gamma 1 and gamma 2 gamma 2 do not obey Michaelis-Menten kinetics at pH 7.5 but exhibit negative cooperativity with Hill coefficients of 0.54 and 0.55 and S0.5 values of 1.0 and 0.63 mM, respectively. However, all isoenzymes display Michaelis-Menten kinetics for ethanol oxidation at pH 10.0 with Km values ranging from 1.5 to 3.2 mM. The maximum specific activity of beta 1 beta 1 is considerably lower than that of the other three homodimers at both pH 7.5 and 10.0. The Km values of the four homodimers for NAD+ at pH 7.5 range from 7.4 to 13 microM and those for NADH, from 6.4 to 33 microM. Ki values for NADH range from 0.19 to 1.6 microM. At pH 7.5, the kinetic properties of alpha alpha and beta 1 beta 1, prepared in vitro from dissociated and recombined alpha beta 1, are similar to those of the native homodimers. The forms gamma 1 gamma 1 and gamma 2 gamma 2, prepared from dissociated and recombined alpha gamma 1 and beta 1 gamma 2, respectively, exhibit negative cooperativity with Hill coefficients that are similar to those seen with the respective native homodimers.
Using magnetic field and plasma observations from four of the five Time History of Events and Macroscale Interactions during Substorms (THEMIS) spacecraft, a surface wave at the dawn flank of the ...magnetopause was identified on 25 April 2007. The wave had an amplitude of about 1 R^sub E^ and propagated tailward with a velocity of about 190-240 km/s. Its azimuthal wavelength was in the range of 9-11 R^sub E^. Magnetosheath velocity values support the hypothesis that this surface wave was generated by the Kelvin-Helmholtz instability. Simultaneously, an ultralow-frequency (ULF) pulsation event was detected by the fifth THEMIS spacecraft deeper in the magnetosphere, at a distance of about 5-7 R^sub E^ from the magnetopause. This ULF event showed all the signatures predicted for waves generated by the classical field line resonance process. Frequency and phases of the detected ULF oscillations were found to be in close agreement with the magnetopause surface periodic disturbances. We conclude that the observed ULF wave event was most likely directly generated by the magnetopause surface wave and thus represents one of the few known manifestations of the classical field line resonance process in space directly observed, a conclusion made possible due to the special configuration of the THEMIS mission and its five spacecraft.
Four alcohol dehydrogenase isoenzymes with "atypical" pH optima for ethanol oxidation at 8.8 were isolated from Japanese livers with the homozygous ADH2 2-2 and the heterozygous ADH2 2-1 phenotypes. ...Agarose gel isoelectric focusing patterns after dissociation--recombination of three isoenzymes purified from the homozygous livers indicate that they are beta 2 beta 2, alpha beta 2, and beta 2 gamma 1. A fourth isoenzyme, purified from livers with the heterozygous phenotype by agarose-hexane--AMP affinity chromatography, was identified as beta 2 beta 1 by dissociation-recombination studies. The kinetic properties of the three heterodimers, beta 2 beta 1, alpha beta 2, and beta 2 gamma 1, are intermediate between those of the respective homodimers, suggesting that the two subunits act independently. Product inhibition studies indicate that beta 2 beta 2 obeys an ordered sequential mechanism, as do the alpha alpha, beta 1 beta 1, gamma 1 gamma 1, and gamma 2 gamma 2 homodimers which have the "typical" pH optimum for ethanol oxidation at pH 10.0-10.5. The kinetic constants of beta 2 beta 2 differ substantially from those of the other homodimers. At pH 7.5, the Vmax for ethanol oxidation of beta 2 beta 2 is 5-40 times higher than that of alpha alpha, beta 1 beta 1, gamma 1 gamma 1, and gamma 2 gamma 2. The Km and Ki values of beta 2 beta 2 for NAD+ and NADH are also considerably higher than those of the other homodimers.
In order to relate the catalytic properties of alcohol dehydrogenase (ADH), the rate-limiting enzyme for alcohol metabolism, with the pharmacokinetics of ethanol elimination in vivo, the multiple ...molecular forms of dog liver ADH were purified and their steady state kinetics investigated. Two different classes of ADH forms were identified by starch gel electrophoresis: the class I isoenzymes migrate to the cathode and the class II forms migrate to the anode. Three different patterns of the cathodic class I isoenzymes were identified in different liver specimens. Three molecular forms were observed for patterns A and C, and five for B. The two classes of isoenzymes were separated by affinity chromatography and purified by column chromatography. The three predominant class I isoenzymes, A1, B2, and C1, in type A, B, and C livers, respectively, were isolated by high performance cation-exchange chromatography. The steady state kinetic constants of the A1, B2, and C1 isoenzymes are similar, but differ substantially from those of the class II enzyme. The class II enzyme is much less sensitive to pyrazole inhibition, Ki = 2 mM, than the class I forms, Ki = 0.6 microM. Methanol is not a substrate for the class II enzyme, whereas it is oxidized by the class I isoenzymes. The class I isoenzymes exhibit a lower Km and substrate inhibition Ki for ethanol, 0.4 and 160 mM, respectively, than values for the class II enzyme, 10 and 610 mM, respectively. The properties of class I and II dog liver ADH are similar to those of the respective isoenzymes purified from human and monkey liver.
The first direct measurements of nuclear tensor polarization p{sub zz} in a laser-driven polarized D target have been performed at Argonne. We present p{sub zz} and electron polarization P{sub e} ...data taken at a magnetic field of 600 G in the optical pumping cell. These results are highly indicative that spin-temperature equilibrium is achieved in the system. To prevent spin relaxation of D and K atoms as well as the molecular recombination of D atoms, the walls of the laser-driven D target are coated with organosilane compounds. We discuss a new coating technique, the 'afterwash', developed at Argonne which has yielded stable atomic fraction results when the coating is exposed to K. We also present new coating techniques for glass and Cu substrates.
Four alcohol dehydrogenase isoenzymes with "atypical" pH optima for ethanol oxidation at 8.8 were isolated from Japanese livers with the homozygous ADH sub(2) 2-2 and the heterozygous ADH sub(2) 2-1 ...phenotypes. Agarose gel isoelectric focusing patterns after dissociation-recombination of three isoenzymes purified from the homozygous livers indicate that they are beta sub(2) beta sub(2), alpha beta sub(2), and beta sub(2) gamma sub(1). A fourth isoenzyme, purified from livers with the heterozygous phenotype by agarose-hexane-AMP affinity chromatography, was identified as beta sub(2) beta sub(1) by dissociation-recombination studies. The kinetic properties of the three heterodimes, beta sub(2) beta sub(1), alpha beta sub(2), and beta sub(2) gamma sub(1), are intermediate between those of the respective homodimers, suggesting that the two subunits act independently.
New molecular forms of human liver alcohol dehydrogenase (ADH), collectively designed ADH Indianapolis (ADHInd), were recently discovered in 29% of liver specimens from Black Americans Bosron, W. F., ...Li, T.-K., and Vallee, B. L. (1981). Proc. Natl. Acad.Sci. USA 77:5784. Three different ADHInd phenotypes have now been identified by starch gel electrophoresis, and four ADHInd enzyme forms isolated by affinity and ion-exchange chromatography. The most cathodic ADHInd form has a single pH optimum at 7.0 for ethanol oxidation and is a homodimer of a newly discovered subunit, as evidenced by dissociation--recombination studies. The remaining three purified ADHInd forms have dual pH optima for ethanol oxidation at 7.0 and 10.0 and generate two new bands on starch gel electrophoresis after dissociation-recombination. They appear to be heterodimers of this new subunit with the known subunits, alpha, beta 1, and gamma 1. Based on the occurrence of these four ADHInd isozymes and isozymes containing beta 1 subunits in the homogenate supernatants of 135 livers, we conclude that ADHInd results from polymorphism at the ADH2 locus, with the variant ADH2Ind allele coding for the beta Ind subunit. The frequency of ADH2Ind was 0.16 in Black Americans. The frequency of the ADH31 and ADH32 alleles also differed in these two populations.
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