Medical Physics Research Today is a free monthly online journal that collates and summarizes the latest research about Medical Physics, including details on medicine, radiotherapy, biomechanics, medical imaging. | ||||||||
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Biophysical Analysis of Progressive C-Terminal Truncations of Human Apolipoprotein E4: Insights into Secondary Structure and Unfolding Properties.Chroni A, Pyrpassopoulos S, Thanassoulas A, Nounesis G, Zannis VI, Stratikos E Institute of Biology, Bio-molecular Physics Laboratory, and Protein Chemistry Laboratory, IRRP, National Centre for Scientific Research "Demokritos", Aghia Paraskevi, Athens 15310, Greece, Molecular Genetics, Departments of Medicine and Biochemistry, Whitaker Cardiovascular Institute, Boston University School of Medicine, Boston, Massachusetts 02118, and Department of Basic Sciences, University of Crete Medical School, Heraklion 71110, Greece achroni@bio.demokritos.gr, stratos@rrp.demokritos.gr. Apolipoprotein E4 (apoE4) is a risk factor for Alzheimer's disease and has been associated with a variety of neuropathological processes. ApoE4 C-terminally truncated forms have been found in brains of Alzheimer's disease patients. Structural rearrangements in apoE4 are known to be key to its physiological functions. To understand the effect of C-terminal truncations on apoE4 lipid-free structure, we produced a series of recombinant apoE4 forms with progressive C-terminal deletions between residues 166 and 299. Circular dichroism measurements show a dramatic loss in helicity upon removal of the last 40 C-terminal residues, whereas further truncations of residues 203-259 lead to recovery of helical content. Further deletion of residues 186-202 leads to a small increase in helical content. Thermal denaturation indicated that removal of residues 260-299 leads to an increase in melting temperature but truncations down to residue 186 did not further affect the melting temperature. The progressive C-terminal truncations, however, gradually increased the cooperativity of thermal unfolding. Chemical denaturation of the apoE4 forms revealed a two-step process with a clear intermediate stage that is progressively lost as the C-terminus is truncated down to residue 230. Hydrophobic fluorescent probe binding suggested that regions 260-299 and 186-202 contain hydrophobic sites, the former being solvent accessible in the wild-type molecule and the latter being accessible only upon truncation. Taken together, our results show an important but complex role of apoE4 C-terminal segments in secondary structure stability and unfolding and suggest that interactions mediated by the C-terminal segments are important for the structural integrity and conformational changes of apoE4. Published 11 August 2008 in Biochemistry.
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