Oxidative metabolism of dopamine: a colour reaction from human midbrain analysed by mass spectrometry.
In order to identify the protein responsible for a dopamine peroxidizing activity, previously described in human normal and parkinsonian substantia nigra by our group, we developed non-denaturing polyacrylamide gel electrophoresis conditions, mimicking the characteristic colour in vitro reaction, resulting from cyclic oxidation of dopamine (DA). After separating protein mixtures from human normal midbrain homogenates on two sets of identical native gels, one gel set was subjected to specific activity staining by using DA and hydrogen peroxide. An activity red/orange band appeared in midbrain tissue lanes, similarly to the lane where commercial horseradish peroxidase (HRP) was present as control of peroxidative activity. The second set of gels, stained with Coomassie Blue, showed other, not enzymatically active protein bands. Mass spectrometry analysis of the bands containing the activity and the corresponding Coomassie Blue bands revealed the presence of proteins that may play a role in neurodegenerative disease, highlighting a possible functional link among dopamine/dopaminochrome redox cycle and protein metabolism.
De Iuliis
Angela
A
Department of Medical Sciences, University of Padua, Padova, Italy. angela.deiuliis@unipd.it
Arrigoni
Giorgio
G
Andersson
Liselotte
L
Zambenedetti
Pamela
P
Burlina
Alessandro
A
James
Peter
P
Arslan
Paola
P
Vianello
Fabio
F
D000368:Aged; D000369:Aged, 80 and over; D003116:Color; D004298:Dopamine / Q000032:analysis / Q000378:metabolism*; D005260:Female; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D008636:Mesencephalon / Q000378:metabolism*; D008954:Models, Biological; D010084:Oxidation-Reduction; D009195:Peroxidase / Q000378:metabolism; D011506:Proteins / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism; D013194:Staining and Labeling / Q000379:methods*; D013379:Substrate Specificity
D011506:Proteins; D006861:Hydrogen Peroxide; D009195:Peroxidase; D004298:Dopamine