Characterization of the residual adenosine deaminating activity in the spleen of a patient with combined immunodeficiency disease and adenosine deaminase deficiency

Abstract

A number of infants with an autosomal recessive form of combined immunodeficiency disease also lack adenosine deaminase (adenosine aminohydrolase; EC 3.5.4.4) activity in their erythrocytes. Other tissues from these infants contain only a few percent of the adenosine-deaminating activity present in corresponding normal tissue. The residual adenosine-deaminating activity in extracts from the spleen of a combined immunodeficient, adenosine deaminase-deficient patient was compared with adenosine deaminase from normal spleen. Affinity and immunoadsorbant column chromatography revealed distinct differences between the adenosine-deaminating activity in the patient's spleen and adenosine deaminase from normal spleen. The point of maximum activity and general configuration of the pH optimum curves were also different. erythro -9-(2-Hydroxyl-3-nonyl)adenine, a potent inhibitor of adenosine deaminase from normal spleen, had relatively little effect on the activity from the patient's spleen. In contrast, adenine was a better inhibitor of the activity in the patient's spleen than it was of the enzyme from normal tissue. An adenosine-deaminating activity with the same characteristics and specific activity as that in the patient's spleen was also isolated from normal spleen. These results suggest that the adenosine-deaminating activity in the spleen of this patient is not due to a mutant form of adenosine deaminase.

Erythrocyte Adenosine Deaminase Deficiency without Immunodeficiency

“…Schrader et al (10) have described an additional ADA activity in some normal tissues which differs from the major ADA in that it is active only at high substrate concentrations, is not inhibitable by EHNA (an inhibitor of the usual form of ADA), and is found in normal amounts in tissues of an ADA-deficient child. In accordance with isozyme nomenclature we have tentatively denoted the high- (Table I vs. Table II).…”

“…To increase the specificity of the ADA assay, enzyme activity was determined at low substrate concentration and as that activity which could be inhibited by 50 jsM EHNA, an inhibitor of ADA. Such assay conditions should eliminate measurement of the activity of a second isozyme of ADA (ADA2) which is present in normal amounts in ADA,--SCID patients, is active at high substrate concentrations, and is not inhibitable by EHNA (10). Additionally, the assays were performed in tris EDTA buffer to inhibit possible formation of inosine by sequential phosphorylation, deamination, and dephosphorylation of adenosine through alternative pathways.…”

Biochemical and Functional Abnormalities in Lymphocytes from an Adenosine Deaminase-deficient Patient during Enzyme Replacement Therapy

“…We examined ADA2 in an extract of human leukemic granulocytes in which ADA2 comprised -2% of total ADA activity. ADA2 was isolated from this extract by Sephadex G-200 chromatography (24). ADA2 had a high estimated Michaelis constant (Ki) (2-5 mM) for both adenosine and deoxyadenosine and utilized deoxyadenosine at 27% of the rate of utilization of adenosine.…”

“…The major isozyme of ADA (ADA1) is inhibited by erythro-9-(2-hydroxy-3-nonyl)-adenine and dCF (23)(24)(25). A minor deaminating activity (ADA2) appears to be the product of a genetic locus separate from ADA1, is not as readily inhibited by erythro-9-(2-hydroxy-3-nonyl)adenine as ADA1 (23)(24)(25), and does not cross-react with antiserum to ADA1 (23,24). The effect of dCF on human ADA2 has not been reported.…”

“…The amounts of residual ADA activity have differed from patient to patient, and the nature of this activity is not always clear. Distinctions among the possible sources of ADA activity can be made using combinations of specific enzyme inhibitors and quantitative immunoassays for protein reacting with antiserum to ADA, (24,26,28,29). Protein antigenically related to ADA, is easily detected in both normal and deficient lymphocytes by radioimmunoassay (Table I).…”