3-nitrotyrosine and Pituitary-Neoplasms

Oxidative stress is extensively associated with tumorigenesis. A series of studies on stable tyrosine nitration as a marker of oxidative damage were performed in human pituitary and adenoma. This paper reviews published research on the mass spectrometry characteristics of nitropeptides and nitroproteomics of pituitary controls and adenomas. The methodology used for nitroproteomics, the current status of human pituitary nitroproteomics studies, and the future perspectives are reviewed. Enrichment of those low-abundance endogenous nitroproteins from human tissues or body fluid samples is the first important step for nitroproteomics studies. Mass spectrometry is the essential approach to determine the amino acid sequence and locate the nitrotyrosine sites. Bioinformatics analyses, including protein domain and motif analyses, are needed to locate the nitrotyrosine site within the corresponding protein domains/motifs. Systems biology techniques, including pathway analysis, are necessary to discover signaling pathway networks involving nitroproteins from the systematically global point of view. Future quantitative nitroproteomics will discover pituitary adenoma-specific nitroprotein(s). Structural biology techniques such as X-ray crystallography analysis will solidly clarify the effects of tyrosine nitration on structure and functions of a protein. Those studies will eventually address the mechanisms and biological functions of tyrosine nitration in pituitary tumorigenesis and will discover nitroprotein biomarkers for pituitary adenomas and targets for drug design for pituitary adenoma therapy.

Topics: Computational Biology; Humans; Metabolic Networks and Pathways; Nitrites; Oxidative Stress; Pituitary Neoplasms; Proteomics; Reactive Nitrogen Species; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Tyrosine

The chemically stable tyrosine nitration of a protein involves the addition of a nitro group (-NO(2)) to the phenolic ring of a tyrosine residue, which may be associated with nervous system physiological and pathological processes. Identification of nitrotyrosine sites on a protein could clarify the functional significance of the modification. Due to the rarity of nitrotyrosine sites in a proteome, tandem mass spectrometry, coupled with different techniques that isolate and enrich nitrotyrosine-containing proteins from a pituitary proteome, is currently the most effective method for site identification. Commercially available nitrotyrosine polyclonal/monoclonal antibodies enable one to detect nitrotyrosine-containing proteins in a two-dimensional gel electrophoresis (2DGE) map, and to preferentially enrich nitrotyrosine-containing proteins with immunoprecipitation. Our present protocols have integrated different isolation/enrichment techniques (2DGE; Western blots; nitrotyrosine immunoaffinity precipitation) and two different tandem mass spectrometry methods (MALDI-MS/MS; ESI-MS/MS) to determine the amino acid sequence of nitrotyrosine-containing peptides that derive from nitrated proteins. Bioinformatics tools are then used to correlate nitrotyrosine sites with a functional domain/motif in order to understand the relationship between tyrosine nitration and the structural/functions of proteins.

Topics: Computational Biology; Electrophoresis, Gel, Two-Dimensional; Humans; Isoelectric Focusing; Mass Spectrometry; Pituitary Neoplasms; Proteins; Proteomics; Tyrosine

The aim of this study was to characterize endogenous nitroproteins, and those proteins that interact with nitroproteins, in a human pituitary nonfunctional adenoma so as to clarify the role of protein nitration in adenomas. A nitrotyrosine affinity column (NTAC) was used to preferentially enrich and isolate endogenous nitroproteins and nitroprotein-protein complexes from a tissue homogenate that was prepared from a human pituitary nonfunctional pituitary adenoma. The preferentially enriched endogenous nitroproteins and nitroprotein-protein complexes were subjected to trypsin digestion, desalination, and tandem mass spectrometry analysis. Nine nitroproteins (Rho-GTPase-activing protein 5, leukocyte immunoglobulin-like receptor subfamily A member 4 precursor, zinc finger protein 432, cAMP-dependent protein kinase type I-beta regulatory subunit, sphingosine-1-phosphate lyase 1, centaurin beta 1, proteasome subunit alpha type 2, interleukin 1 family member 6, and rhophilin 2) and three proteins (interleukin 1 receptor-associated kinase-like 2, glutamate receptor-interacting protein 2, and ubiquitin) that interacted with nitroproteins were discovered. The nitration site of each nitroprotein was located onto the functional domain where nitration occurred, and each nitroprotein was related to a corresponding functional system. Those data indicate that protein nitration might be an important molecular event in the formation of a human pituitary nonfunctional adenoma.

Topics: Adenoma; Adult; Amino Acid Sequence; Chromatography, Affinity; Humans; Male; Molecular Sequence Data; Multiprotein Complexes; Neoplasm Proteins; Nitrogen; Pituitary Neoplasms; Protein Structure, Tertiary; Proteomics; Sensitivity and Specificity; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Trypsin; Tyrosine