Detection and characterization of variant and modified structures of proteins in blood and tissues by mass spectrometry

Shimizu, Akira; Nakanishi, Toyofumi; Miyazaki, Akira

doi:10.1002/mas.20086

Cited by 43 publications

(46 citation statements)

References 109 publications

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“…9 Purified fibrinogen from synovial fluids of RA patients was identified by Western blot analysis with anticitrulline antibody as described above. Bands were developed with ECL Plus.…”

Section: Immunoprecipitation and Western Blot With Anticitrulline Antmentioning

confidence: 99%

Detection of citrullinated proteins in synovial fluids derived from patients with rheumatoid arthritis by proteomics-based analysis

Tabushi¹,

Nakanishi²,

Takeuchi³

et al. 2008

Ann Clin Biochem

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Background: Rheumatoid arthritis (RA) is the most common inflammatory joint disease. The aetiology of RA remains unknown, but autoimmune responses are considered to play an important role in the disease pathophysiology. Currently available data suggests that the process of diagnosing RA may benefit from testing for anticyclic citrullinated peptides. Identification of the presence of citrullinated proteins in rheumatoid synovial fluids is important for the elucidation of the aetiology of RA as well as in the differential diagnosis of rheumatic-related diseases. Methods: A proteomics-based approach using electrophoresis/mass spectrometry was applied to identify the citrullinated proteins in synovial fluids from patients with RA. Synovial fluids from patients with RA were subjected to sodium dodecyl sulfate -polyacrylamide gel electrophoresis and Western blot analysis to detect the citrullinated proteins. Identification bands were then subjected to mass spectrometry. Results: Three proteins -citrullinated fibrinogen, citrullinated fibronectin and citrullinated vimentin -in synovial fluids from RA patients were identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. Conclusions: Proteomics-based analysis can be used to detect citrullinated proteins in synovial fluids from RA patients.

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“…9 Purified fibrinogen from synovial fluids of RA patients was identified by Western blot analysis with anticitrulline antibody as described above. Bands were developed with ECL Plus.…”

Section: Immunoprecipitation and Western Blot With Anticitrulline Antmentioning

confidence: 99%

Detection of citrullinated proteins in synovial fluids derived from patients with rheumatoid arthritis by proteomics-based analysis

Tabushi¹,

Nakanishi²,

Takeuchi³

et al. 2008

Ann Clin Biochem

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“…ESI-MS has been widely used to detect Hb variants in hemoglobin [21][22][23][24][25] and to investigate its structural assembly into a noncovalent complex [26 -29] and its corresponding disassembly [30].…”

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confidence: 99%

Probing hemoglobin structure by means of traveling-wave ion mobility mass spectrometry

Scarff

Patel

Thalassinos

et al. 2008

J. Am. Soc. Mass Spectrom.

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Hemoglobin (Hb) is a tetrameric noncovalent complex consisting of two ␣-and two ␣ ␤-globin ␤ chains each associated with a heme group. Its exact assembly pathway is a matter of debate. Disorders of hemoglobin are the most common inherited disorders and subsequently the molecule has been extensively studied. This work attempts to further elucidate the structural properties of the hemoglobin tetramer and its components. Gas-phase conformations of hemoglobin tetramers and their constituents were investigated by means of traveling-wave ion mobility mass spectrometry. Sickle (HbS) and normal (HbA) hemoglobin molecules were analyzed to determine whether conformational differences in their quaternary structure could be observed. Rotationally averaged collision cross sections were estimated for tetramer, dimer, apo-, and holo-monomers with reference to a protein standard with known cross sections. Estimates of cross section obtained for the tetramers were compared to values calculated from X-ray crystallographic structures. HbS was consistently estimated to have a larger cross section than that of HbA, comparable with values obtained from X-ray crystallographic structures. Nontetrameric species observed included apo-and holo-forms of ␣-and ␣ ␤-monomers and ␤ heterodimers; ␣-and ␣ ␤-monomers in both apo-and holo-forms were found to have similar ␤ cross sections, suggesting they maintain a similar fold in the gas phase in both the presence and the absence of heme. Heme-deficient dimer, observed in the spectrum when analyzing commercially prepared Hb, was not observed when analyzing fresh blood. This implies that holo-␣-apo-␣ ␤ is not an essential intermediate within the The typical electrospray ionization (ESI) mass spectrum of a protein consists of an envelope of peaks attributed to a series of multiply charged gas-phase ions that can indicate the stability and compactness of its structure in the gas phase [4,5]. Multiply charged ions are produced by proton attachment, predominantly to exposed basic sites on the protein [6], and those of lowest charge are thought to be most representative of native structure [7]. A highly compact protein would have fewer exposed basic sites than those of an unfolded conformation of the same protein and thus would accept fewer charges [4,8].Ion mobility spectrometry is a shape-selective technique, based on the time taken for an ion to traverse a mobility cell containing an inert gas under the influence of a weak electric field [9]. This time is related to the rotationally averaged collision cross section, mass, and charge of the ion. The coupling of ion mobility separation with mass spectrometry has provided a powerful method for the analysis of complex mixtures and for the determination of molecular structure [10].Ion mobility mass spectrometry has emerged as a complementary technique to the well-established methods of X-ray crystallography and nuclear magnetic resonance (NMR) spectroscopy for three-dimensional protein structure analysis [11]. There is now substantial evidence to support...

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“…Furthermore, variants with no clinical manifestations could cause substantial overestimation of HbA 1c , the important marker for diabetes mellitus. Worldwide, an estimated 150 million people carry Hb variants (3 ), and hemoglobinopathies are the commonest inherited disorders, constituting a significant healthcare problem (4 ). Therefore, reliable detection and identification methods are essential.…”

mentioning

confidence: 99%

Mass Spectrometry: A Tool for Enhanced Detection of Hemoglobin Variants

Kleinert¹,

Schmid

Zurbriggen

et al. 2008

Clinical Chemistry

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Detection and characterization of variant and modified structures of proteins in blood and tissues by mass spectrometry

Cited by 43 publications

References 109 publications

Detection of citrullinated proteins in synovial fluids derived from patients with rheumatoid arthritis by proteomics-based analysis

Detection of citrullinated proteins in synovial fluids derived from patients with rheumatoid arthritis by proteomics-based analysis

Probing hemoglobin structure by means of traveling-wave ion mobility mass spectrometry

Mass Spectrometry: A Tool for Enhanced Detection of Hemoglobin Variants

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