Deciphering structural intermediates and genotoxic fibrillar aggregates of albumins: a molecular mechanism underlying for degenerative diseases.
The misfolding and aggregation of proteins is involved in some of the most prevalent neurodegenerative disorders. The importance of human serum albumin (HSA) stems from the fact that it is involved in bio-regulatory and transport phenomena. Here the effect of acetonitrile (ACN) on the conformational stability of HSA and by comparison, ovalbumin (OVA) has been evaluated in the presence and absence of NaCl. The results show the presence of significant amount of secondary structure in HSA at 70% ACN and in OVA at 50% ACN, as evident from far-UV Circular Dichroism (CD) and Attenuated Total Reflection Fourier transformed infra red spectroscopy (ATR-FTIR). Tryptophan and 8-Anilino-1-Naphthalene-Sulphonic acid (ANS) fluorescence indicate altered tryptophan environment and high ANS binding suggesting a compact "molten globule"-like conformation with enhanced exposure of hydrophobic surface area. However, in presence of NaCl no intermediate state was observed. Detection of aggregates in HSA and OVA was possible at 90% ACN. Aggregates possess extensive β-sheet structure as revealed by far-UV CD and ATR-FTIR. These aggregates exhibit increase Thioflavin T (Th T) fluorescence with a red shift of Congo red (CR) absorption spectrum. X-ray diffraction (XRD) and Scanning Electron Microscopy (SEM) analysis confirmed the presence of fibrillar aggregates. Single cell gel electrophoresis (SCGE) assay of these fibrillar aggregates showed the DNA damage resulting in cell necrosis confirming their genotoxic nature. Some proteins not related to any human disease form fibrils in vitro. In the present study ACN gives access to a model system to study the process of aggregation.
Department of Biochemistry, Faculty of Life Science, Aligarh Muslim University, Aligarh, Uttar Pradesh, India. aabgeenanaim@gmail.com
Aabgeena
A
Naeem
Samreen
S
Amani
D000097:Acetonitriles / Q000494:pharmacology; D000418:Albumins / Q000737:chemistry*; D052160:Benzothiazoles; D002942:Circular Dichroism; D003224:Congo Red / Q000737:chemistry; D006801:Humans; D008214:Lymphocytes / Q000187:drug effects / Q000378:metabolism; D008855:Microscopy, Electron, Scanning; D019636:Neurodegenerative Diseases / Q000378:metabolism*; D010047:Ovalbumin / Q000737:chemistry; D017433:Protein Structure, Secondary; D012965:Sodium Chloride / Q000494:pharmacology; D017550:Spectroscopy, Fourier Transform Infrared; D013844:Thiazoles / Q000737:chemistry; D014961:X-Ray Diffraction
D000097:Acetonitriles; D000418:Albumins; D052160:Benzothiazoles; D013844:Thiazoles; C009462:thioflavin T; D003224:Congo Red; D012965:Sodium Chloride; D010047:Ovalbumin; C032159:acetonitrile