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Determination of a Histidine Residue at the Yeast Orotate Phosphoribosyltransferase Active-Site

  • Donald L. Sloan
  • Rosalyn S. Strauss
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 165)

Abstract

A purified preparation of orotate phosphoribosyltransferase(0-PRTase) from yeast has been shown to catalyze the formation of orotidine monophosphate (OMP) from orotate and 5-phosphoribosyl α-1-pyrophosphate (PRibPP) through the use of a ping pong kinetic mechanism (1) in the presence of an optimum concentration of Mgll(2). The present investigation was initiated to search for nucleo-philic active-site residues of 0-PRTase which would stabilize (in an SN1 elimination) or react with (in a triple SN2 displacement) the enzyme-phosphoribosyl intermediate, formed as a consequence of this mechanism. Analysis of the amino acid composition, pH dependency of the 0-PRTase activity in the forward and reverse directions, and chemical modification of 0-PRTase using histidine-specific reagents have been accomplished.

Keywords

Amino Acid Composition Histidine Residue Enzyme Active Site Orotate Phosphoribosyltransferase Active Site Histidine 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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References

  1. 1.
    J. Victor, L B. Greenberg, and D. L. Sloan, Studies of the Kinetic Mechanism of Orotate Phosphoribosyltransferase from Yeast, J. Biol. Chem. 254: 2647 (1979).PubMedGoogle Scholar
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    J. Victor, A. Leo-Mensah, and D. L. Sloan, Divalent Metal Ion Activation of the Yeast Orotate Phosphoribosyltransferase Catalyzed Reaction, Biochemistry 18: 3597 (1979).PubMedCrossRefGoogle Scholar
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    S. Stein, P. Bohlem, J. Stone, W. Dairman, and S. Udenfriend, Amino Acid Analysis with Fluorescamine at the Picomole Level, Arch. Biochem. Biophys. 155: 202 (1973).PubMedCrossRefGoogle Scholar
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    G. W. Miles, Modification of Histidine Residues in Proteins by Diethylpyrocarbonate, Meth. Enzymol. 47: 431 (1977).PubMedCrossRefGoogle Scholar
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    K. F. Tipton and H. B. F. Dixon, Effects of pH on Enzymes, Meth. Enzymol. 63: 183 (1979).CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 1984

Authors and Affiliations

  • Donald L. Sloan
    • 1
  • Rosalyn S. Strauss
    • 1
  1. 1.Chemistry DeptCity College of the City University of New YorkNew YorkUSA

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