Oxidative modification of ovalbumin.
Stimulated neutrophils (PMNL) are a source of the active oxygen species: O2, H2O2 and HOCl/OCl- which in turn can act on proteins yielding a variety of mixed oxidation products. A system is proposed in which a model protein-ovalbumin (OVA) first undergoes chlorination by HOCl/OCl- and next is oxidised by H2O2. The modification of functional groups (-NH2, -SH, -S-S-, > C = O, Tyr and Trp) in OVA was monitored as well as their accessibility to promote aggregation. Chlorination resulted in additional inter- or intra -S-S- bond formation followed by a decrease in the total sulfhydryl group content. Amino groups were oxidised to carbonyl moieties with a concomitant acidic shift of pI. Formation of chlorotyrosine at the chlorination step was confirmed and its further H2O2-mediated transformation to bityrosine was demonstrated. It has also been confirmed that tryptophan, and not tyrosine, is the first target for chlorination. SDS/PAGE and HPLC profiles revealed that HOCl/OCl- chlorination promotes formation of aggregates stabilised by non covalent bonds. In conclusion, we suggest that a dramatic change in the OVA molecule structure begins when the molar excess of HOCl/OCl- is about 2 per one reactive group in OVA.
Olszowski
S
S
Jagiellonian University, Collegium Medicum, Institute of Medical Biochemistry, Cracow, Poland.
Olszowska
E
E
Stelmaszyńska
T
T
Krawczyk
A
A
Marcinkiewicz
J
J
Baczek
N
N
D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D006997:Hypochlorous Acid / Q000737:chemistry; D007525:Isoelectric Focusing; D010047:Ovalbumin / Q000737:chemistry*; D010084:Oxidation-Reduction; D013438:Sulfhydryl Compounds / Q000737:chemistry
D013438:Sulfhydryl Compounds; D006997:Hypochlorous Acid; D010047:Ovalbumin