Mechanistic studies on uroporphyrin I-induced photoinactivation of some heme-enzymes.
Aerobic and anaerobic studies have demonstrated that uroporphyrin I-induced inactivation of delta-aminolevulinic acid dehydratase, porphobilinogenase, deaminase and uroporphyrinogen decarboxylase was dependent on oxygen and mediated by reactive oxygen species. The mechanism of photoinactivation of those heme-enzymes from human erythrocytes by uroporphyrin I by u.v. light was investigated. Enzymes of the heme pathway were preincubated in the presence of specific scavengers for several reactive oxygen species and then exposed to uroporphyrin I and u.v. light. Upon exposure of the enzymes to the porphyrin under u.v. light, and in an aerobic atmosphere, the percentage of enzyme activities with respect to the corresponding controls were 50.2 +/- 5.1 (SD, n = 6), 25.3 +/- 3.0 (SD, n = 6), 25.9 +/- 2.8 (SD, n = 6) and 49.7 +/- 7.5 (SD, n = 8) for delta-aminolevulinic acid dehydratase, porphobilinogenase, deaminase and uroporphyrinogen decarboxylase, respectively. The presence of sodium azide, histidine or superoxide dismutase did not protect the enzymes against the effects of uroporphyrin I. However, both cysteine and potassium ferrycyanide prevented the enzyme photoinactivation induced by uroporphyrin I. In the presence of either catalase or GSH, the enzyme photoinactivation was lower. Ethanol, glucose and dimethylsulfoxide had no effect on enzyme activity, while ion chelators had variable effects. This study shows that the type II mechanism is not the predominant reaction mediating the uroporphyrin I effect and enzyme photoinactivation would involve an electron transfer. Hydrogen peroxide and hydroxyl radicals could possibly mediate the uroporphyrin I-induced enzyme photoinactivation.
Afonso
S G
SG
Centro de Investigaciones sobre Porfirinas y Porfirias (CIPYP), University of Buenos Aires, Argentina.
Polo
C F
CF
Enriquez de Salamanca
R
R
Batlle
A
A
D000642:Ammonia-Lyases / Q000528:radiation effects; D004583:Electrons; D016166:Free Radical Scavengers; D006420:Hemeproteins / Q000528:radiation effects*; D006801:Humans; D006861:Hydrogen Peroxide / Q000097:blood; D017665:Hydroxyl Radical; D011163:Hydroxymethylbilane Synthase / Q000528:radiation effects; D008190:Lyases / Q000528:radiation effects*; D010100:Oxygen / Q000097:blood; D000623:Porphobilinogen Synthase / Q000528:radiation effects; D013481:Superoxides / Q000097:blood; D014575:Uroporphyrinogen Decarboxylase / Q000528:radiation effects; D014578:Uroporphyrins / Q000494:pharmacology*
D016166:Free Radical Scavengers; D006420:Hemeproteins; D014578:Uroporphyrins; D013481:Superoxides; D017665:Hydroxyl Radical; C034103:uroporphyrin I; D006861:Hydrogen Peroxide; D011163:Hydroxymethylbilane Synthase; D008190:Lyases; D014575:Uroporphyrinogen Decarboxylase; D000623:Porphobilinogen Synthase; D000642:Ammonia-Lyases; C100132:porphobilinogenase; D010100:Oxygen