TY - JOUR
T1 - Center of Mass Calculation in Combination with MS/MS Allows Robust Identification of Single Amino Acid Polymorphisms in Clinical Measurements of Insulin-Like Growth Factor-1
AU - Maus, Anthony
AU - Kemp, Jennifer
AU - Milosevic, Dragana
AU - Renuse, Santosh
AU - Pandey, Akhilesh
AU - Singh, Ravinder J.
AU - Grebe, Stefan K.G.
N1 - Publisher Copyright:
Copyright © 2019 American Chemical Society.
PY - 2020/1/3
Y1 - 2020/1/3
N2 - Insulin-like growth factor-1 (IGF-1) measurement by high-resolution accurate mass-mass spectrometry (HRAM-MS) is replacing IGF-1 immunoassays and allows for identification of single amino acid variants; by contrast, both normal and deleterious sequence variants might be missed by immunoassays or non-HRAM-MS methods. We have developed an intact molecule HRAM-MS method to identify IGF-1 variants, distinguishing them by a center of mass (COM) calculation, followed by various tandem-MS activation techniques (HCD, ETD, ETciD, EThcD, UVPD). We found single amino acid variants in 841 of 146 620 patient samples (0.57%). Most were benign (A67T, A70T). We also observed a pathogenic variant (V44M), likely pathogenic variants (A38V, V17M), and a likely benign variant (A67V). For 207 samples from unique patients with residual serum, the MS variant results were confirmed by cell-free DNA sequencing. Our approach allows accurate quantitative reporting of functional IGF-1 in the presence of single amino acid variants. The COM approach potentially enables omission of tandem-MS for known, common variants, while the combination of COM and tandem-MS allows accurate identification in all cases we encountered. This approach should be applicable to qualitative and quantitative analyses of other peptides/proteins in clinical and research settings and might lend itself to the characterization of other protein variations.
AB - Insulin-like growth factor-1 (IGF-1) measurement by high-resolution accurate mass-mass spectrometry (HRAM-MS) is replacing IGF-1 immunoassays and allows for identification of single amino acid variants; by contrast, both normal and deleterious sequence variants might be missed by immunoassays or non-HRAM-MS methods. We have developed an intact molecule HRAM-MS method to identify IGF-1 variants, distinguishing them by a center of mass (COM) calculation, followed by various tandem-MS activation techniques (HCD, ETD, ETciD, EThcD, UVPD). We found single amino acid variants in 841 of 146 620 patient samples (0.57%). Most were benign (A67T, A70T). We also observed a pathogenic variant (V44M), likely pathogenic variants (A38V, V17M), and a likely benign variant (A67V). For 207 samples from unique patients with residual serum, the MS variant results were confirmed by cell-free DNA sequencing. Our approach allows accurate quantitative reporting of functional IGF-1 in the presence of single amino acid variants. The COM approach potentially enables omission of tandem-MS for known, common variants, while the combination of COM and tandem-MS allows accurate identification in all cases we encountered. This approach should be applicable to qualitative and quantitative analyses of other peptides/proteins in clinical and research settings and might lend itself to the characterization of other protein variations.
KW - center of mass calculation
KW - high-resolution accurate mass-MS
KW - insulin-like growth factor-1
KW - intact molecule MS
KW - single amino acid protein variants
KW - tandem mass spectrometry
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U2 - 10.1021/acs.jproteome.9b00494
DO - 10.1021/acs.jproteome.9b00494
M3 - Article
C2 - 31736316
AN - SCOPUS:85076539974
SN - 1535-3893
VL - 19
SP - 186
EP - 193
JO - Journal of Proteome Research
JF - Journal of Proteome Research
IS - 1
ER -