ECTO-enzyme activity of human erythrocyte adenosine deaminase

  • Ken Bielat
  • George L. Tritsch
Original Article

DOI: 10.1007/BF00222613

Cite this article as:
Bielat, K. & Tritsch, G.L. Mol Cell Biochem (1989) 86: 135. doi:10.1007/BF00222613

Summary

Adenosine deaminase is found primarily in the cytoplasm of many cell types. In the human erythrocyte, about 30 per cent of the total adenosine deaminase activity is membrane associated, and about two-thirds of this is inactivated by treatment of intact erythrocytes with the nonpenetrating reagent diazotized sulfanilic acid, without affecting lactate dehydrogenase, a soluble cytoplasmic enzyme. This indicates that within the cell membranes, the catalytic site of about two-thirds of the adenosine deaminase faces the external medium, i.e., ecto adenosine deaminase. Localization of adenosine deaminase activity at the cell membrane is demonstrated directly by electron microscopy by use of the substrate 6-Chloropurine ribonucleoside, which is dechlorinated by adenosine deaminase to produce Cl, which is precipitated at its locus of formation by added Ag+, and the precipitated AgCl converted into the electron dense Ag0 upon exposure to light.

From the Hydropathic Profile of the amino acid sequence of adenosine deaminase it is evident that there are two hydrophobic domains of sufficient length to span a biological membrane, and it is proposed that these domains could function to anchor the enzyme to the membrane.

The importance of adenosine deaminase is indicated by the fatal immuno-deficiency which results from untreated genetic adenosine deaminase deficiency. It may be important to determine whether the amount of ecto adenosine deaminase activity is better suited to assess the clinical status of adenosine deaminase deficient patients that the currently used total cellular enzyme activity.

Key words

ecto-enzymeadenosine deaminaseerythrocyteelectron microscopy

Abbreviations

ADA

Adenosine Deaminase

LDH

Lactate Dehydrogenase

HEPES

N-2-Hydroxyethylpiperazine-N′-2-ethanesulfonic acid

CPR

6-Chloropurine Ribonucleoside

SDS

Sodium Dodecyl Sulfate

NAD

β-Nicotinamide Adenine Dinucleotide

HBSS

Hank's Balanced Salt Solution

DASA

Diazotized Sulfanilic Acid

Copyright information

© Kluwer Academic Publishers 1989

Authors and Affiliations

  • Ken Bielat
    • 1
  • George L. Tritsch
    • 1
  1. 1.Roswell Park Memorial InstituteBuffaloUSA