PMID: 40614Nov 9, 1979

Characterization of the active site of homogeneous thyroid purine nucleoside phosphorylase

Biochimica Et Biophysica Acta
J D Carlson, A G Fischer

Abstract

Purine nucleoside phosphorylase (purine-nucleoside : orthophosphate ribosyltransferase, EC 2.4.2.1) has been purified approx. 4000-fold and to electrophoretic homogeneity from bovine thyroid glands. The isolated enzyme has a specific activity of 17 mumol . min-1 . mg-1. The native enzyme appears to have a molecular weight of 92 000 as determined by sedimentation equilibrum ultracentrifugation and is comprised of three subunits having a molecular weight of 31 000 each as shown by sodium dodecyl sulfate gel electrophoresis. The enzyme is irreversibly denatured below pH 5 and the enzyme-substrate complex is shown to have an ionization constant (pKa) of 9.2 which influences catalytic activity. The pH dependence of the kinetic constants identifies three amino acid ionizable protons. The binding of inosine is effected by an imidazole ring of histidine (pKa 5.65) and a sulfhydryl group of cysteine (pKa 8.5) and the maximal velocity is restricted by an epsilon-amino group which is essential for phosphate binding. The requirement of these residues for activity was confirmed by group-specific chemical modification. The presence of phosphate protected only the lysyl residue while inosine protected all three residues from chemical titratio...Continue Reading

References

Jun 1, 1975·Archives of Biochemistry and Biophysics·T P Moyer, A G Fischer
May 1, 1977·Analytical Biochemistry·J J Sedmak, S E Grossberg
Oct 1, 1978·Archives of Biochemistry and Biophysics·F Jordan, A Wu
Sep 1, 1967·Acta paediatrica Scandinavica·K S LeeK Berg
Mar 1, 1964·Biochemistry·D A YPHANTIS
Apr 1, 1964·Archives of Biochemistry and Biophysics·M I SIMON, H VANVUNAKIS

Citations

Jan 1, 1980·The International Journal of Biochemistry·T P MoyerA R Schulz
Dec 18, 2001·The Biochemical Journal·Matthew J Pugmire, Steven E Ealick
Jun 15, 1991·Biochemical Pharmacology·A BzowskaN G Johansson

Related Concepts

Metazoa
Bos indicus
Hydrogen-Ion Concentration
Models, Structural
Pentosyltransferase
Nicotinamide Riboside Phosphorylase
Thyroid Gland

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