Rapid Purification and High Sensitivity Analysis of Heparan Sulfate from Cells and Tissues TOWARD GLYCOMICS PROFILING

Guimond SE, Puvirajesinghe TM, Skidmore MA, Kalus I, Dierks T, Yates EA, Turnbull JE (2009)
JOURNAL OF BIOLOGICAL CHEMISTRY 284(38): 25714-25722.

Zeitschriftenaufsatz | Veröffentlicht | Englisch
 
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Abstract / Bemerkung
Studies on glycosaminoglycans and proteoglycans (PGs) have been hampered by difficulties in isolation and analysis by traditional methods that are laborious and lack sensitivity and throughput. Here we demonstrate a simple method for rapid isolation of proteoglycans (RIP) employing phenol/guanidine/chloroform reagent to purify heparan sulfate (HS) PGs quantitatively from various tissues and cells. We further show that this generic purification methodology, when applied in concert with a BODIPY (TM) fluorescent label, permits structural analyses on RIP-purified HS at similar to 1,000-fold higher sensitivity than standard UV detection methods and similar to 10-100-fold higher sensitivity than previous fluorescence detection methods. The utility of RIP-BODIPY methodology was demonstrated by rapid profiling of HS structural composition from small tissue samples, multiple mouse organs, and as little as a few thousand cultured cells. It was also used to generate novel insights into in vivo structural changes in HS from Sulf1 knock-out mice for the first time that differed significantly from previous observations limited to tissue culture experiments. RIP was also applied to purify HS for bioassay testing, exemplified by cell assays of fibroblast growth factor signaling activation; this generated data from 2-O-sulfotransferase knock-out mice and revealed an unexpected deficiency in fibroblast growth factor activation by HS from heterozygous mice. These data demonstrate that RIP will underpin emerging efforts to develop glycomics profiling strategies for HS and other glycosaminoglycans to explore their structure-function relationships in complex biological systems.
Erscheinungsjahr
2009
Zeitschriftentitel
JOURNAL OF BIOLOGICAL CHEMISTRY
Band
284
Ausgabe
38
Seite(n)
25714-25722
ISSN
0021-9258
eISSN
1083-351X
Page URI
https://pub.uni-bielefeld.de/record/1590897

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Guimond SE, Puvirajesinghe TM, Skidmore MA, et al. Rapid Purification and High Sensitivity Analysis of Heparan Sulfate from Cells and Tissues TOWARD GLYCOMICS PROFILING. JOURNAL OF BIOLOGICAL CHEMISTRY. 2009;284(38):25714-25722.
Guimond, S. E., Puvirajesinghe, T. M., Skidmore, M. A., Kalus, I., Dierks, T., Yates, E. A., & Turnbull, J. E. (2009). Rapid Purification and High Sensitivity Analysis of Heparan Sulfate from Cells and Tissues TOWARD GLYCOMICS PROFILING. JOURNAL OF BIOLOGICAL CHEMISTRY, 284(38), 25714-25722. doi:10.1074/jbc.M109.032755
Guimond, S. E., Puvirajesinghe, T. M., Skidmore, M. A., Kalus, I., Dierks, T., Yates, E. A., and Turnbull, J. E. (2009). Rapid Purification and High Sensitivity Analysis of Heparan Sulfate from Cells and Tissues TOWARD GLYCOMICS PROFILING. JOURNAL OF BIOLOGICAL CHEMISTRY 284, 25714-25722.
Guimond, S.E., et al., 2009. Rapid Purification and High Sensitivity Analysis of Heparan Sulfate from Cells and Tissues TOWARD GLYCOMICS PROFILING. JOURNAL OF BIOLOGICAL CHEMISTRY, 284(38), p 25714-25722.
S.E. Guimond, et al., “Rapid Purification and High Sensitivity Analysis of Heparan Sulfate from Cells and Tissues TOWARD GLYCOMICS PROFILING”, JOURNAL OF BIOLOGICAL CHEMISTRY, vol. 284, 2009, pp. 25714-25722.
Guimond, S.E., Puvirajesinghe, T.M., Skidmore, M.A., Kalus, I., Dierks, T., Yates, E.A., Turnbull, J.E.: Rapid Purification and High Sensitivity Analysis of Heparan Sulfate from Cells and Tissues TOWARD GLYCOMICS PROFILING. JOURNAL OF BIOLOGICAL CHEMISTRY. 284, 25714-25722 (2009).
Guimond, Scott E., Puvirajesinghe, Tania M., Skidmore, Mark A., Kalus, Ina, Dierks, Thomas, Yates, Edwin A., and Turnbull, Jeremy E. “Rapid Purification and High Sensitivity Analysis of Heparan Sulfate from Cells and Tissues TOWARD GLYCOMICS PROFILING”. JOURNAL OF BIOLOGICAL CHEMISTRY 284.38 (2009): 25714-25722.

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