Mass Spectrometric Phenotyping of Val34Leu Polymorphism of Blood Coagulation Factor XIII by Differential Peptide Display

Abstract

Background:The Val34Leu mutation in the activation peptide of factor XIII (FXIIIA) correlates with a lower incidence of myocardial infarction and ischemic stroke but an increased risk for hemorrhagic stroke. We describe mass spectrometric detection of the activation peptide variants in human serum. Methods: We used differential peptide display (DPD) to compare comprehensive peptide maps from pairs of serum samples from healthy volunteers. Peptides were separated by liquid chromatography, and fractions were subjected to mass spectrometry. Mass spectra of all fractions were combined, giving a peptide map representing a two-dimensional display of peptide masses. After comparison of peptide mass maps, peptides that differentiated FXIIIA phenotypes were identified by mass spectrometry. Results: Val34Leu polymorphisms of the activation peptide of FXIIIA were identified in 20 serum samples from 10 volunteers by DPD, and their sequences were confirmed by nanoelectrospray-ionization quadrupole time-of-flight mass spectrometry. Analysis of three (V34V, V34L, and L34L) phenotypes was confirmed by allele-specific genotypic analysis in all (n ‫؍‬ 10) volunteers. Conclusion: DPD provides a simple and easy-to-use phenotype assay with advantages over PCR-based assays in being faster and directly analyzing the compound of interest.

Serum protein profiling in mice: Identification of Factor XIIIa as a potential biomarker for muscular dystrophy

“…Macrophages are known to infiltrate dystrophic muscle [29], which may explain the lower level of this protein in serum. The same peptide has also previously been identified by mass spectrometry in human serum where a Val34Leu mutation of the protein is correlated with a lower incidence of myocardial infarction and ischemic stroke [30], where cardiovascular abnormalities are well characterized in the mdx mouse [31]. Finally, in two separate gene expression studies, expression of FXIIIa1 was found to be 6-to 10-fold higher in differentiating compared to undifferentiated human control myo-…”

Search for Breast Cancer Biomarkers in Fractionated Serum Samples by Protein Profiling With SELDI‐TOF MS

“…Based on their m/z , the sample type and fraction in which they were found, the ProteinChip surface used, as well as data from previously performed serum profiling studies (N. Harris, unpublished data, and ), we tentatively identified the most discriminative peaks. The peak with m/z 15,915 that was detected in Fraction 3 on the IMAC30 array is very likely hemoglobin β‐chain.…”

Processing and classification of protein mass spectra

“…Similar to microarray or electrophoretic approaches, two LC-MS maps from different samples can be color coded as red and blue images and overlaid. The resulting image shows upregulated or downregulated peptides as red or blue, respectively, and the unchanged peptides as dark magenta (Tammen et al, 2004). To make this approach work, the two LC-MS runs have to be well aligned.…”