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Deciphering aggregates, prefibrillar oligomers and protofibrils of cytochrome c

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Abstract

Aggregation of protein into insoluble intracellular complexes and inclusion bodies underlies the pathogenesis of human neurodegenerative diseases. Importance of cytochrome c (cyt c) arises from its involvement in apoptosis, sequence homology and for studying molecular evolution. A systemic investigation of polyethylene glycol (PEG) and trifluoroethanol (TFE) on the conformational stability of cyt c as a model hemeprotein was made using multi-methodological approach. Cyt c exists as molten globule (MG) at 60 % PEG-400 and 40 % TFE as confirmed by far-UV CD, attenuated total reflection Fourier transform infrared spectroscopy, Trp environment, 8-anilino-1-naphthalene-sulfonic acid (ANS) binding and blue shift in the soret band. Q-band splitting in MG states specifies conformational changes in the hydrophobic heme-binding pocket. Aggregates were detected at 90 % PEG-400 and 50 % TFE as confirmed by increase thioflavin T and ANS fluorescence and shift in Congo red absorbance. Detection of prefibrils and protofibrils at 90 % PEG-400 and 50 % TFE was possible after 72-h incubation. Single cell gel electrophoresis of prefibrils and protofibrils showed DNA damage confirming their toxicity and potential health hazards. Scanning electron microscopy and XRD analysis confirmed prefibrillar oligomers and protofibrils of cyt c.

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Acknowledgments

The authors are highly thankful for the facilities available at AMU Aligarh. Authors are thankful to Indo–US Science and Technology Forum for financial support. USIF, AMU has been highly acknowledged for providing SEM facility. S. A. is the recipient of UGC-SRF.

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The authors declare that there are no conflicts of interest.

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Authors and Affiliations

  1. Department of Biochemistry, Faculty of Life Science, Aligarh Muslim University, Aligarh, 202002, India

    Samreen Amani & Aabgeena Naeem

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  1. Samreen Amani
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  2. Aabgeena Naeem
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Correspondence to Aabgeena Naeem.

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Amani, S., Naeem, A. Deciphering aggregates, prefibrillar oligomers and protofibrils of cytochrome c. Amino Acids 46, 1839–1851 (2014). https://doi.org/10.1007/s00726-014-1698-y

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  • DOI: https://doi.org/10.1007/s00726-014-1698-y

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