The influence of heme-binding proteins in heme-catalyzed oxidations.
We show here that heme-binding proteins may enhance, decrease, or completely inhibit heme-catalyzed oxidations and that in doing so the proteins themselves may be oxidized depending upon their relative affinities for heme and the nature of their interactions with this metalloporphyrin. That release of iron from heme was not responsible for the catalytic effect is indicated by the observation that heme induced more peroxidation of rat liver microsomal lipid in the presence of H2O2 than iron and that iron release is very low under the conditions employed. Hemopexin, which binds heme with high affinity, completely inhibited heme-catalyzed lipid peroxidation at concentrations slightly higher than that of heme, suggesting a unique role for this acute phase protein in antioxidant defense mechanisms. The protein itself was not oxidized, presumably because the putative bis-histidyl heme-hemopexin complex cannot interact with H2O2. Rat and human albumin and rat glutathione S-transferases (GST), proteins with moderate affinities for heme, decreased heme-catalyzed lipid peroxidation in a dose-dependent manner but were subject to oxidation. The GST were crosslinked forming a nondisulfide covalently linked subunit dimer as well as products of higher molecular weight whereas the oxidation products of the albumins had molecular weights only slightly higher than those of the native proteins. The changes in the electrophoretic patterns of GST and albumin were accompanied by a decrease in their tryptophan fluorescence and the formation of bityrosine-like products. Proteins with lower affinities for heme, such as bovine albumin and rat liver fatty acid-binding protein (L-FABP), enhanced lipid peroxidation at all concentrations tested. While bovine albumin was modified, L-FABP was not crosslinked nor were its tyrosine residues oxidized. Thus, the susceptibility of a protein to heme-mediated oxidative damage would appear to be determined by factors such as its affinity for heme, the nature of the amino acids in the vicinity of the bound catalyst and the availability of a free coordination site on the iron.
Vincent
S H
SH
Department of Pediatrics, Cornell University Medical College, New York, New York 10021.
Grady
R W
RW
Shaklai
N
N
Snider
J M
JM
Muller-Eberhard
U
U
D000818:Animals; D002352:Carrier Proteins / Q000494:pharmacology*; D002712:Chlorides; D004591:Electrophoresis, Polyacrylamide Gel; D005290:Ferric Compounds / Q000494:pharmacology; D006418:Heme / Q000494:pharmacology*; D000081062:Heme-Binding Proteins; D006420:Hemeproteins; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007501:Iron / Q000378:metabolism; D007502:Iron Chelating Agents / Q000494:pharmacology; D008054:Lipid Peroxides / Q000096:biosynthesis*; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism*; D010084:Oxidation-Reduction / Q000187:drug effects; D011485:Protein Binding; D051381:Rats; D013050:Spectrometry, Fluorescence
D002352:Carrier Proteins; D002712:Chlorides; D005290:Ferric Compounds; D000081062:Heme-Binding Proteins; D006420:Hemeproteins; D007502:Iron Chelating Agents; D008054:Lipid Peroxides; D006418:Heme; D006861:Hydrogen Peroxide; D007501:Iron; C024555:ferric chloride