The smoke-borne electrophile acrolein reacts extensively with proteins, forming carbonyl-retaining Michael adducts that may be attacked by adjacent protein nucleophiles to form cross-links. Because little information is available concerning the extent of intermolecular protein cross-linking during acrolein toxicity in cells, we used an antibody against a known target for toxic carbonyls, the chaperone protein Hsp90, to detect the formation of high-mass protein complexes in acrolein-exposed A549 cells. A 3 h exposure to acrolein (0 to 200 µM) resulted in concentration-dependent formation of a single high-mass band (approx. 180 kDa). This species was detected in cells exposed to just 50 µM acrolein, a concentration that did not elicit acute cell death as assessed by measurements of cell ATP levels. The formation of cross-linked Hsp90 coincided with a rapid loss of carbonyl adducts within cells that had been subjected to a brief “pulse” exposure to a subtoxic concentration of acrolein, suggesting Michael adducts are short-lived within cells due in part to consumption during reactions with protein nucleophiles. Cross-linked Hsp90 persisted following an overnight recovery incubation, suggesting the cellular ability to repair or degrade these species is limited. Two known carbonyl scavengers, hydralazine and bisulfite, strongly protected against the ATP depletion accompanying acrolein exposure, but only the latter suppressed protein adduction and Hsp90 cross-linking. As previously shown for hydralazine, mass spectrometry studies using a model peptide indicated that bisulfite traps carbonyl groups possessed by Michael addition adducts, and such adduct-trapping reactivity appeared to contribute to the blockade of Hsp90 cross-linking in acrolein-preloaded cells. Collectively, these findings establish that formation of stable intermolecular protein cross-links accompanies exposure to acrolein. Future clarification of the chemistry underlying this damage may provide novel biomarkers of acrolein exposure.

EXPLOR_STEP=$WICRI_ROOT/Sante/explor/ChloroquineV1/Data/Main/Exploration

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{{Explor lien
   |wiki=    Sante
   |area=    ChloroquineV1
   |flux=    Main
   |étape=   Exploration
   |type=    RBID
   |clé=     ISTEX:45204FA66D2197D9BB4DF82B5AC171BC67A207D5
   |texte=   Intermolecular Protein Cross-Linking During Acrolein Toxicity: Efficacy of Carbonyl Scavengers as Inhibitors of Heat Shock Protein-90 Cross-Linking in A549 Cells
}}