LC-MS/MS Analysis Unravels Deep Oxidation of Manganese Superoxide Dismutase in Kidney Cancer.
Manganese superoxide dismutase (MNSOD) is one of the major scavengers of reactive oxygen species (ROS) in mitochondria with pivotal regulatory role in ischemic disorders, inflammation and cancer. Here we report oxidative modification of MNSOD in human renal cell carcinoma (RCC) by the shotgun method using data-dependent liquid chromatography tandem mass spectrometry (LC-MS/MS). While 5816 and 5571 proteins were identified in cancer and adjacent tissues, respectively, 208 proteins were found to be up- or down-regulated (p< 0.05). Ontological category, interaction network and Western blotting suggested a close correlation between RCC-mediated proteins and oxidoreductases such as MNSOD. Markedly, oxidative modifications of MNSOD were identified at histidine (H54and H55), tyrosine (Y58), tryptophan (W147, W149, W205and W210) and asparagine (N206and N209) residues additional to methionine. These oxidative insults were located at three hotspots near the hydrophobic pocket of the manganese binding site, of which the oxidation of Y58, W147and W149was up-regulated around three folds and the oxidation of H54and H55was detected in the cancer tissues only (p< 0.05). When normalized to MNSOD expression levels, relative MNSOD enzymatic activity was decreased in cancer tissues, suggesting impairment of MNSOD enzymatic activity in kidney cancer due to modifications. Thus, LC-MS/MS analysis revealed multiple oxidative modifications of MNSOD at different amino acid residues that might mediate the regulation of the superoxide radicals, mitochondrial ROS scavenging and MNSOD activity in kidney cancer.
Cancer Research Center, Shandong University School of Medicine, Jinan 250012, China. zhaozuohui@126.com.|Departments of Urology and Pediatric Surgery, Shandong Provincial Hospital Qianfoshan Hospital, Shandong University, No. 324, Jingwu Road, Jinan 250021, China. zhaozuohui@126.com.
Zuohui
Z
Zhao
Departments of Surgery, Urology, Pathology and Proteomics Laboratory, Veterans Affairs Boston Healthcare System, Boston University School of Medicine, 150 S. Huntington Ave., Boston, MA 02130, USA. kazadzoi@bu.edu.
Kazem M
KM
Azadzoi
Departments of Surgery, Urology, Pathology and Proteomics Laboratory, Veterans Affairs Boston Healthcare System, Boston University School of Medicine, 150 S. Huntington Ave., Boston, MA 02130, USA. hpchoi89@gmail.com.
Han-Pil
HP
Choi
Cancer Research Center, Shandong University School of Medicine, Jinan 250012, China. jingruirui223@163.com.
Ruirui
R
Jing
Cancer Research Center, Shandong University School of Medicine, Jinan 250012, China. lvxin121@163.com.
Xin
X
Lu
Cancer Research Center, Shandong University School of Medicine, Jinan 250012, China. cuilingli@yahoo.com.
Cuiling
C
Li
Cancer Research Center, Shandong University School of Medicine, Jinan 250012, China. wfqlzq@163.com.
Fengqin
F
Wang
Departments of Urology and Pediatric Surgery, Shandong Provincial Hospital Qianfoshan Hospital, Shandong University, No. 324, Jingwu Road, Jinan 250021, China. kyoto2310@sina.com.
Jiaju
J
Lu
Cancer Research Center, Shandong University School of Medicine, Jinan 250012, China. jyang@bu.edu.|Departments of Surgery, Urology, Pathology and Proteomics Laboratory, Veterans Affairs Boston Healthcare System, Boston University School of Medicine, 150 S. Huntington Ave., Boston, MA 02130, USA. jyang@bu.edu.
Jing-Hua
JH
Yang
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D002853:Chromatography, Liquid; D016000:Cluster Analysis; D004789:Enzyme Activation; D006801:Humans; D007680:Kidney Neoplasms / Q000235:genetics / Q000378:metabolism*; D055432:Metabolomics / Q000379:methods; D008958:Models, Molecular; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D011485:Protein Binding; D011487:Protein Conformation; D025941:Protein Interaction Mapping; D060066:Protein Interaction Maps; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction; D013482:Superoxide Dismutase / Q000737:chemistry / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
D000596:Amino Acids; D017382:Reactive Oxygen Species; D013482:Superoxide Dismutase
MNSOD; ROS; liquid chromatography-tandem mass spectrometry; oxidation; quantitative proteomics