Organocatalytic removal of formaldehyde adducts from RNA and DNA bases.
Saswata Karmakar, Emily M Harcourt, David S Hewings, Florian Scherer, Alexander F Lovejoy, David M Kurtz, Thomas Ehrenschwender, Luzi J Barandun, Caroline Roost, Ash A Alizadeh, Eric T Kool
Index: Nature Chemistry 7 , 752-8, (2015)
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Abstract
Formaldehyde is universally used to fix tissue specimens, where it forms hemiaminal and aminal adducts with biomolecules, hindering the ability to retrieve molecular information. Common methods for removing these adducts involve extended heating, which can cause extensive degradation of nucleic acids, particularly RNA. Here, we show that water-soluble bifunctional catalysts (anthranilates and phosphanilates) speed the reversal of formaldehyde adducts of mononucleotides over standard buffers. Studies with formaldehyde-treated RNA oligonucleotides show that the catalysts enhance adduct removal, restoring unmodified RNA at 37 °C even when extensively modified, while avoiding the high temperatures that promote RNA degradation. Experiments with formalin-fixed, paraffin-embedded cell samples show that the catalysis is compatible with common RNA extraction protocols, with detectable RNA yields increased by 1.5-2.4-fold using a catalyst under optimized conditions and by 7-25-fold compared with a commercial kit. Such catalytic strategies show promise for general use in reversing formaldehyde adducts in clinical specimens.
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