Detection, Extraction And Identification Of Typical Proteins Of VSD Serum

Objective: to detect, extract and identify VSD-specific serum proteins and study the association between their physiological functions and cardiac development.Methods : (1). screening of serum marker proteins of VSD: Serum samples were collected from 30 VSD patients and from 30 normal controls. The serum protein expression patterns of the two groups were examined through surface-enhanced laser desorption and ionization time of flight mass spectrometry (SELDI-TOF-MS) and differential proteins were further analysed. Data were processed by Biomarker Wizard and Biomarker Parrern. (2). Abstracting and determining of VSD-marker serum proteins:Differential proteins were separated in polyacrylamide gel electrophoresis (PAGE) and were further abstracted and analysed by LC-MS. Bioinformatic methods were applied to analyse target proteins and to illustrate their structures and functions.Results: (1). VSD-marker serum proteins: a). there are 15 proteins statistically significant (P<0.05) from the VSD group having differential expressions from those of the normal control group (molecular weight ranging from 2000 to 20000Da); six of them have up-regulated expressions and nine of them have down-regulated expressions. b). The protein with molecular weight of 15141Da and 15884Da from the VSD group shows the most significant differential expression from its counterpart in the normal control group. (The ratio between the compared peaks is five.) (2). Abstracting and determining of VSD-marker serum proteins: a). A significantly differential protein band is separated by PAGE from VSD group at around 15KDa. b). Four unique protein chains were detected of Profilin-1 through LTQ Orbitrap mass spectrum identification, with reliability of 100% and coverage of 93%. Six unique protein chains were detected of CystatinC through LTQ Orbitrap mass spectrum identification, with reliability of 100% and coverage of 90%. c). Bioinformatic analysis shows that Profilin-1 contains 140 amino acids. It's molecular weight is 15054Da, which is identical with that of the target protein detected through SELDI. The four unique protein chains detected are completely matching with human Profilin-1. CystatinC contains 146 amino acids. It's molecular weight is 15779Da, which is identical with that of the target protein detected through SELDI. The 6 unique protein chains detected are completely matching with human CystatinC.Conclusions: (1). There are discrepancy between serum protein expression patterns of VSD patients and those of the normal controls. (2). Proteins with molecular weights of 5419Da,5640Da,7779Da,15141 Da and 15884Da may be differential proteins relative to VSD. (3). Profilin-1 and CystatinC may be one of the VSD-marker serum proteins. (4). SELDI-TOF-MS and LTQ-Orbitrap mass spectrum identification can be new approaches to searching for and determining biomarkers of diseases.