High resolution mass spectrometric alveolar proteomics: identification of surfactant protein SP-A and SP-D modifications in proteinosis and cystic fibrosis patients.
In the present study, one- and two-dimensional gel electrophoresis combined with high resolution Fourier transform-ion cyclotron resonance mass spectrometry (FT-ICR MS) have been applied as powerful approaches for the proteome analysis of surfactant proteins SP-A and SP-D, including identification of structurally modified and truncation forms, in bronchoalveolar lavage fluid from patients with cystic fibrosis, chronic bronchitis and pulmonary alveolar proteinosis. Highly sensitive micropreparation techniques were developed for matrix-assisted laser desorption/ionization (MALDI) FT-ICR MS analysis which provided the identification of surfactant proteins at very low levels. Owing to the high resolution, FT-ICR MS was found to provide substantial advantages for the structural identification of surfactant proteins from complex biological matrices with high mass determination accuracy. Several protein bands corresponding to SP-A and SP-D were identified by MALDI-FT-ICR MS after electrophoretic separation by one- and two-dimensional gel electrophoresis, and provided the identification of structural modifications (hydroxy-proline) and degradation products. The high resolution mass spectrometric proteome analysis should facilitate the unequivocal identification of subunits, aggregations, modifications and degradation products of surfactant proteins and hence contribute to the understanding of the mechanistic basis of lung disease pathogenesis.
Laboratory of Analytical Chemistry, Department of Chemistry, University of Konstanz, Konstanz, Germany.
Yu
Y
Bai
Dmitry
D
Galetskiy
Eugen
E
Damoc
Christian
C
Paschen
Zhiqiang
Z
Liu
Mathias
M
Griese
Shuying
S
Liu
Michael
M
Przybylski
D001992:Bronchoalveolar Lavage Fluid / Q000737:chemistry; D003550:Cystic Fibrosis / Q000503:physiopathology*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D040901:Proteomics / Q000379:methods*; D037662:Pulmonary Surfactant-Associated Protein A / Q000032:analysis*; D037663:Pulmonary Surfactant-Associated Protein D / Q000032:analysis*
D037662:Pulmonary Surfactant-Associated Protein A; D037663:Pulmonary Surfactant-Associated Protein D