Transferrin variants: Pitfalls in the diagnostics of Congenital disorders of glycosylation

Zühlsdorf, Andrea; Park, Julien H.; Wada, Yoshinao; Rust, Stephan; DuChesne, Ingrid; Grüneberg, Marianne; Marquardt, Thorsten

doi:10.1016/j.clinbiochem.2014.09.022

Cited by 32 publications

(22 citation statements)

References 11 publications

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“…Transferrin analysis: Tf IEF, HPLC Refs. 5,12,17 and Tf neuraminidase digestion 6,9 were performed according to standard methods.…”

Section: Methodsmentioning

confidence: 99%

“…5,6 A type 1 pattern points to a glycan assembly defect (CDG-I; cytosol and ER), a type 2 pattern to a glycan remodeling defect (CDG-II; Golgi or vesicular defect). [6][7][8][9] An abnormal pattern must be checked in a new serum sample for confirmation. Before considering genetic testing, a secondary CDG, such as galactosemia and fructose intolerance, has to be excluded.…”

Section: Introductionmentioning

confidence: 99%

“…Before considering genetic testing, a secondary CDG, such as galactosemia and fructose intolerance, has to be excluded. 9,10 Genetic testing is often performed by analyzing a CDG gene panel. If this is normal, whole exome or whole genome sequencing should be the next step.…”

Section: Introductionmentioning

confidence: 99%

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Ten years of screening for congenital disorders of glycosylation in Argentina: case studies and pitfalls

et al. 2018

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BACKGROUND: Congenital Disorders of Glycosylation (CDG) are genetic diseases caused by hypoglycosylation of glycoproteins and glycolipids. Most CDG are multisystem disorders with mild to severe involvement. METHODS: We studied 554 patients (2007-2017) with a clinical phenotype compatible with a CDG. Screening was performed by serum transferrin isoelectric focusing. The diagnosis was confirmed by genetic testing (Sanger or exome sequencing). RESULTS: A confirmed abnormal pattern was found in nine patients. Seven patients showed a type 1 pattern: four with PMM2-CDG, two with ALG2-CDG, and one with classical galactosemia. A type 2 pattern was found in two patients: one with a CDG-IIx and one with a transferrin protein variant. Abnormal transferrin pattern were observed in a patient with a myopathy due to a COL6A2 gene variant. CONCLUSIONS: CDG screening in Argentina from 2007 to 2017 revealed 4 PMM2-CDG patients, 2 ALG2-CDG patients with a novel homozygous gene variant and 1 CDG-IIx.

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“…Transferrin analysis: Tf IEF, HPLC Refs. 5,12,17 and Tf neuraminidase digestion 6,9 were performed according to standard methods.…”

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Ten years of screening for congenital disorders of glycosylation in Argentina: case studies and pitfalls

et al. 2018

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“…And, certainly, as it has been extensively proved in the last years, MS is the most reliable way to obtain structural information about protein glycosylation as it allows fast and high sensitivity profiling and accurate characterization of heterogeneous glycan structures . Additionally, Tf natural variants may lead to erroneous conclusions when analysing Tf glycoforms by the routine methods described before due to a shift in the bands or peaks, which can be completely avoided using MS . The analysis of Tf glycoforms by MS can be carried out by analysing the intact glycoprotein or by analysing the glycans or glycopeptides obtained by enzymatic digestion .…”

Section: Resultsmentioning

confidence: 99%

Improved tryptic digestion assisted with an acid-labile anionic surfactant for the separation and characterization of glycopeptide glycoforms of a proteolytic-resistant glycoprotein by capillary electrophoresis time-of-flight mass spectrometry

et al. 2015

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Certain glycoproteins are rather difficult to digest due to compacted tertiary or quaternary structures. In a previous study, a capillary LC coupled to TOF-MS (μLC-TOF-MS) method was developed for the detection and characterization of the glycopeptide glycoforms of human transferrin (Tf), a proteolytic resistant glycoprotein, in serum samples. After immunoaffinity purification, Tf was digested with trypsin in the presence of RapiGest(®) and μLC-TOF-MS analyses permitted to detect the N413 and N611 glycopeptide glycoforms. Conversely, the use of this surfactant, albeit mandatory to quantitatively digest the isolated Tf, proved detrimental to CE-TOF-MS analysis due to its interaction with the inner surface of the silica capillary walls. As CE is usually regarded as an interesting alternative to other separation techniques (low consumption of reagents, excellent separation efficiency, and reduced analysis times), in this work, the undesirable interferences of the surfactant have been removed to allow the correct separation and detection of Tf glycoforms by CE-TOF-MS. Moreover, the digestion protocol described by the RapiGest(®) manufacturer has been modified to minimize desialylation of Tf glycopeptide glycoforms. The new developed CE-TOF-MS methodology has been then compared with the former μLC-TOF-MS by means of sensitivity and separation efficiency of Tf glycopeptide glycoforms in the standard glycoprotein. Additionally, Tf glycopeptide glycoforms from serum of healthy volunteers and patients with congenital disorders of glycosylation have also been analyzed following the developed methodology.

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“…Diagnosis of CDG is challenging in the presence of transferrin variants (1)(2)(3). The neuraminidase removes charged sialic acid residues from the transferrin, revealing any changes in the isoelectric point of the transferrin molecule (2)(3).…”

Section: Answersmentioning

confidence: 99%

What's Wrong with the Transferrin?

et al. 2016

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Transferrin variants: Pitfalls in the diagnostics of Congenital disorders of glycosylation

Cited by 32 publications

References 11 publications

Ten years of screening for congenital disorders of glycosylation in Argentina: case studies and pitfalls

Ten years of screening for congenital disorders of glycosylation in Argentina: case studies and pitfalls

Improved tryptic digestion assisted with an acid-labile anionic surfactant for the separation and characterization of glycopeptide glycoforms of a proteolytic-resistant glycoprotein by capillary electrophoresis time-of-flight mass spectrometry

What's Wrong with the Transferrin?

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