Tyrosine modification is not required for myeloperoxidase-induced loss of apolipoprotein A-I functional activities.
Apolipoprotein A-I (apoAI), the major protein of high density lipoprotein, plays an important role in reverse cholesterol transport via its activity as an ABCA1-dependent acceptor of cellular cholesterol. We reported recently that myeloperoxidase (MPO) modification of apoAI inhibits its ABCA1-dependent cholesterol acceptor activity (Zheng, L., Nukuna, B., Brennan, M. L., Sun, M., Goormastic, M., Settle, M., Schmitt, D., Fu, X., Thomson, L., Fox, P. L., Ischiropoulos, H., Smith, J. D., Kinter, M., and Hazen, S. L. (2004) J. Clin. Invest. 114, 529-541). We also reported that MPO-mediated chlorination preferentially modifies two of the seven tyrosines in apoAI, and loss of parent peptides containing these residues dose-dependently correlates with loss in ABCA1-mediated cholesterol acceptor activity (Zheng, L., Settle, M., Brubaker, G., Schmitt, D., Hazen, S. L., Smith, J. D., and Kinter, M. (2005) J. Biol. Chem. 280, 38-47). To determine whether oxidative modification of apoA-I tyrosine residues was responsible for the MPO-mediated inactivation of cholesterol acceptor activity, we made recombinant apoAI with site-specific substitutions of all seven tyrosine residues to phenylalanine. ApoAI and the tyrosine-free apoAI were equally susceptible to dose-dependent MPO-mediated loss of ABCA1-dependent cholesterol acceptor activity, as well as lipid binding activity. MPO modification altered the migration of apoAI on SDS gels and decreased its alpha-helix content. MPO-induced modification also targeted apoAI tryptophan and lysine residues. Specifically, we detected apoAI tryptophan oxidation to mono- and dihydroxytryptophan and apoAI lysine modification to chlorolysine and 2-aminoadipic acid. Thus, tyrosine modification of apoAI is not required for its MPO-mediated inhibition of cholesterol acceptor activity.
Peng
Dao-Quan
DQ
Department of Cell Biology, The Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
Wu
Zhiping
Z
Brubaker
Gregory
G
Zheng
Lemin
L
Settle
Megan
M
Gross
Eitan
E
Kinter
Michael
M
Hazen
Stanley L
SL
Smith
Jonathan D
JD
D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D016632:Apolipoprotein A-I / Q000378:metabolism* / Q000502:physiology; D002784:Cholesterol / Q000378:metabolism*; D018076:DNA, Complementary; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D013058:Mass Spectrometry; D010084:Oxidation-Reduction; D009195:Peroxidase / Q000378:metabolism*; D017354:Point Mutation; D014364:Tryptophan / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism*
C577836:ABCA1 protein, human; D064286:ATP Binding Cassette Transporter 1; D018528:ATP-Binding Cassette Transporters; D016632:Apolipoprotein A-I; D018076:DNA, Complementary; D014443:Tyrosine; D014364:Tryptophan; D002784:Cholesterol; D009195:Peroxidase