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J. Biochem, 1982, Vol. 91, No. 1 167-176
© 1982 Japanese Biochemical Society

research-article

Guanine Deaminase from Rat Brain. Purification, Characteristics, and Contribution to Ammoniagenesis in the Brain

Syuzo MIYAMOTO, Hirofumi OGAWA1, Hiroshi SHIRAKI and Hachiro NAKAGAWA2

Division of Protein Metabolism, Institute for Protein Research, Osaka University Suita, Osaka 565

2 To whom reprint requests should be sent

  1. Guanine deaminase [EC 3.5.4.3 [EC] [EC] ] was purified to a homogeneous state from rat brain by a procedure involving ammonium sulfate fractionation, DE-52 column chromatography, hydroxylapatite column chromatography, gel filtration on ACA-34 and isoelectric focusing. Homogeneity was shown by polyacrylamide gel electro phoresis in the presence and absence of SDS.
  2. The molecular weight of the enzyme was determined by gel filtration (105,000), and that of its subunit by SDS-polyacrylamide gel electrophoresis (52,000). From these findings, we concluded that the native enzyme consisted of two identical subunits.
  3. The Km values for guanine and 8-azaguanine were calculated to be 0.17 mM and 0.67 mM respectively. This enzyme was markedly inhibited by 5-amino 4-imidazolecarboxamide (AICA), a precursor of purine nucleotide synthesis, with aK1 value of 82 µM.
  4. Guanine deaminase was purified from rat liver by the procedure used for purifi cation of the brain enzyme and anti-liver enzyme serum was raised in rabbits. This antiserum cross-reacted with the brain enzyme without spur formation in the Ouchterlony double diffusion test.
  5. The {gamma}-globulin fraction of the anti-guanine deaminase serum and AICA inhibited more than 50% of the ammoniagenesis in the brain system in which the purine nucleotide cycle operates. It was also shown that guanine nucleotides were degraded via guanosine and guanine liberating ammonia in the same brain system when sub strates for the purine nucleotide cycle were omitted.

On the basis of these findings it is suggested that guanine deaminase contributes to ammoniagenesis in the brain.

1 Present address: Department of Biochemistry, Medical School, Toyama Medical and Pharmaceutical University, Sugitani, Toyama 930–01.


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