Characterization of a 5'-nucleotidase and a high affinity Ca2+-stimulated ATPase from microsomes from Zea mays coleoptiles

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1983
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Carter, Stephen
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Biochemistry, Biophysics and Molecular Biology

The Department of Biochemistry, Biophysics, and Molecular Biology was founded to give students an understanding of life principles through the understanding of chemical and physical principles. Among these principles are frontiers of biotechnology such as metabolic networking, the structure of hormones and proteins, genomics, and the like.

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The Department of Biochemistry and Biophysics was founded in 1959, and was administered by the College of Sciences and Humanities (later, College of Liberal Arts & Sciences). In 1979 it became co-administered by the Department of Agriculture (later, College of Agriculture and Life Sciences). In 1998 its name changed to the Department of Biochemistry, Biophysics, and Molecular Biology.

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1959–present

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  • Department of Biochemistry and Biophysics (1959–1998)

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This dissertation is divided into three parts: (1) Purification and Characterization of a 5'-Nucleotidase from Corn Shoot Microsomes, (2) Characterization of a High Affinity (Ca('2+) + Mg('2+))-ATPase from Corn Shoot Plasma Membranes, and (3) Partial Purification and Characterization of a Ca('2+)-Stimulated ATPase Activity from Corn Shoot Plasma Membranes. In the first part, a 5'-nucleotidase was purified to apparent homogeneity. The native enzyme is a glycoprotein, M(,r)= 49,000. SDS/PAGE indicates the presence of two subunits, M(,r) = 24,500 and 25,500. Relative rates of hydrolysis for substrates are: AMP > GMP > IMP > CMP > UMP >> pNPP. Adenosine is a noncompetitive inhibitor, K(,i) UTP > GTP > CTP >> pNPP. The order of the ability of different divalent cations to stimulate the enzyme is Ca('2+) > Mg ('2+) > Co('2+) > Mn('2+) > Ni('2+) > Zn('2+). The enzyme has a K(,a) value for Ca('2+) of 0.2 (mu)M and shows a dependence on Mg('2+) for the high affinity Ca('2+)-ATPase activity to be functional. Part 3 describes the solubilization and partial purification and characterization of a Ca('2+)-ATPase from corn plasma membranes. The enzyme has an apparent molecular weight of 105,000 by gel filtration and a pH optimum at 6.5. The order of the ability of substrates tested to elicit Ca('2+) stimulation by the enzyme is ATP > UTP > CTP > GTP >> pNPP. The order of different divalent cations to stimulate the enzyme is Ca('2+) > Mg('2+) > Co('2+) > Mn('2+) > Ni('2+) > Zn('2+). The Ca('2+) affinity constant determination indicated the presence of two binding affinities by the enzyme, a high affinity component, K(,a) = 0.06 (mu)M and a low affinity component, K(,a) = 15 (mu)M. The addition of monovalent cations increased the velocity of the reaction while not affecting the affinity constant for Ca('2+) with the enzyme; however, monovalent cations were shown to be required for the low affinity component to be functional.

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Sat Jan 01 00:00:00 UTC 1983