Identification of specific protein carbonylation sites in model oxidations of human serum albumin.
Human serum albumin (HSA) was subjected to oxidative stress and the locations of the resulting protein carbonyls were determined using mass spectrometry in conjunction with a hydrazide labeling scheme. To model oxidative stress, HSA samples were subjected to metal-catalyzed oxidation (MCO) conditions or treated with hypochlorous acid (HOCl). Oxidation led to the conversion of lysine residues to 2-aminoadipic semi-aldehyde residues, which were subsequently labeled with biotin hydrazide. Analysis of the tryptic peptides from the samples indicates that the oxidations are highly selective. Under MCO conditions, only two of the 59 lysine residues appeared to be modified (Lys-97 and Lys-186). With HOCl, five different lysine modification sites were identified (Lys-130, Lys-257, Lys-438, Lys-499, and Lys-598). These results strongly suggest that the preferred site of modification is dependent on the nature of the oxidant and that the process relies on specific structural motifs in the protein to direct the oxidation. The high selectivity seen here provides insights into the factors that in vivo drive the selective carbonylation of specific proteins in systems under oxidative stress.
Temple
Ani
A
Department of Chemistry and Biochemistry, San Francisco State University, San Francisco, California, USA.
Yen
Ten-Yang
TY
Gronert
Scott
S
D001665:Binding Sites; D002244:Carbon / Q000737:chemistry*; D006801:Humans; D013058:Mass Spectrometry; D008956:Models, Chemical; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010100:Oxygen / Q000737:chemistry*; D011485:Protein Binding; D012709:Serum Albumin / Q000737:chemistry*; D013329:Structure-Activity Relationship
D012709:Serum Albumin; D002244:Carbon; D010100:Oxygen