Fragile X mental retardation protein is an
mRNA-
binding protein associated with phenotypic manifestations of
fragile X syndrome, an X-linked disorder caused by mutation in the FMR1 gene that is the most common inherited cause of
intellectual disability. Despite the well-studied genetic mechanism of the disease, the proteoforms of
fragile X mental retardation protein have not been thoroughly characterized. Here, we report the expression and mass spectrometric characterization of human
fragile X mental retardation protein. FMR1
cDNA clone was transfected into human HEK293 cells to express the full-length human
fragile X mental retardation protein. Purified
fragile X mental retardation protein was subjected to
trypsin digestion and characterized by mass spectrometry. Results show 80.5%
protein sequence coverage of
fragile X mental retardation protein (Q06787, FMR1_HUMAN) including both the N- and C-terminal
peptides, indicating successful expression of the full-length
protein. Identified post-translational modifications include N-terminal acetylation, phosphorylation (Ser600), and methylation (Arg290, 471, and 474). In addition to the full-length
fragile X mental retardation protein isoform (
isoform 6), two endogenous
fragile X mental retardation protein alternative splicing
isoforms (
isoforms 4 and 7), as well as
fragile X mental retardation protein interacting
proteins, were also identified in the co-purified samples, suggesting the interaction network of the human
fragile X mental retardation protein. Quantification was performed at the
peptide level, and this information provides important reference for the future development of a targeted assay for quantifying
fragile X mental retardation protein in clinical samples. Collectively, this study provides the first comprehensive report of human
fragile X mental retardation protein proteoforms and may help advance the mechanistic understanding of
fragile X syndrome and related phenotypes associated with the FMR1 mutation.