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Rapid Method to Characterize Mutations in Transthyretin in Cerebrospinal Fluid from Familial Amyloidotic Polyneuropathy Patients by Use of Matrix-assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry

¹ Institute of Clinical Neuroscience, Department of Psychiatry and Neurochemistry, and
² Department of Neurology, Göteborg University, Sahlgrenska University Hospital, SE-43/#80 Mölndal, Sweden.

³ Institute of Chemistry, Department of Analytical Chemistry, Uppsala University, PO Box 531, SE-751 21 Uppsala, Sweden.
^a Author for correspondence. Fax 46-18-471-3692; e-mail jonas.bergquist{at}kemi.uu.se

Background: Familial amyloidotic polyneuropathy (FAP) type I, the most common dominantly inherited form of amyloidosis, is caused by a Val-to-Met point mutation at position 30 (Val³⁰Met) in the protein transthyretin. Mass spectrometric analysis can identify modification of proteins, such as point mutations, acetylation, phosphorylation, sulfation, oxidation, and glycosylation.

Methods: Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) spectra from cerebrospinal fluid (CSF) drawn from a patient with FAP were compared with CSF from controls. We also isolated transthyretin with a Centrisart molecular size cutoff filter and performed high-accuracy peptide mass mapping to localize the site of the amino acid substitution (Val³⁰Met).

Results: Mass spectra of transthyretin were produced directly from human CSF as well as from CSF after a simple prepurification method without immunoprecipitation. On-target tryptic digestion and MALDI-MS verified mass spectrometric peak identification. The point mutation was still detectable in CSF after hepatic transplantation.

Conclusions: It is possible to diagnose FAP by a rapid MALDI-TOF MS analysis using only 100 µL of CSF, with only 250 nL actually consumed on target. The approach may also be useful to monitor production of mutated transthyretin by choroid plexus, especially after liver transplantation.

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				D. de Boer, M. M. Erps, W. K.W.H. Wodzig, and M. P. van Dieijen-Visser Inadequate Attempts to Measure the Microheterogeneity of Transthyretin by Low-Resolution Mass Spectrometry Clin. Chem., July 1, 2005; 51(7): 1299 - 1300. [Full Text] [PDF]

				H. R. Bergen III, S. R. Zeldenrust, M. L. Butz, D. S. Snow, P. J. Dyck, P. J. B. Dyck, C. J. Klein, J. F. O'Brien, S. N. Thibodeau, and D. C. Muddiman Identification of Transthyretin Variants by Sequential Proteomic and Genomic Analysis Clin. Chem., September 1, 2004; 50(9): 1544 - 1552. [Abstract] [Full Text] [PDF]