IMMUNOLOGICAL, ENZYMATIC AND BIOCHEMICAL-STUDIES OF UROPORPHYRINOGEN-III SYNTHASE DEFICIENCY IN 20 PATIENTS WITH CONGENITAL ERYTHROPOIETIC PORPHYRIA

Congenital erythropoietic porphyria (CEP), a rare autosomal recessive inborn error of heme biosynthesis, results from reduced activity of ur oporphyrinogen III synthase (URO-III-S) leading to an excessive produc tion and accumulation of porphyrins, Various clinical and biochemical observations point to a relationship between degree of disease express ion and metabolic disturbance. We investigated 20 patients with early onset of clinical symptoms of CEP and, additionally, the relatives of six patients. CEP was confirmed by porphyrinemia and porphyrinuria wit h dominance of uroporphyrin and its isomer I. The investigation of the immunological nature of the defective URO-III-S gene from unrelated p atients with unknown mutations was possible thanks to an antibody agai nst the human enzyme. URO-III-S concentration in erythrocytes was dete rmined by ELISA. No signal was achieved when assaying nonimmune serum by ELISA, whereas there was a positive reaction with the serum after i mmunisation. Furthermore, specificity of immune sera is demonstrated b y immunoprecipitation of URO-III-S activity which caused a 33 % reduct ion of enzyme activity. Normal levels of immunoreactive enzyme protein 100 +/- 10% of control ((x) over bar +/- SD, n = 12) with a reduced s pecific activity 15 +/- 8.5 % ((x) over bar +/- SD, n = 12) were found in erythrocytes from all patients, with the exception of a girl with a remarkably high enzyme concentration of 149% of controls and a very low specific activity of 4%. In consequence, all patients had cross-re acting immunological material (CRIM)-positive mutations. GRIM-ratios v aried between 3.2 and 24.5. The GRIM-positive nature of the gene defec t indicated that the mutations altered the activity of URO-III-S. The different GRIM ratios implied the presence of various mutations, which is further evidence for the heterogeneity in the genetic defect found in CEP, URO-III-S activity was determined in erythrocyte lysates by a coupled enzyme assay. Erythrocyte URO-III-S activities of patients we re reduced to 4-33 % of the normal value (1.72 +/- 0.14 pkat/mg protei n). An increase of urinary coproporphyrin isomer I (40-61%, norm = 17- 31%) and a halved URO-III-S activity can serve as a biochemical test f or asymptomatic heterozygous gene carriers of CEP.