Jing-song Fan1, Honzhen Goh1, Ke Ding2, Bo Xue2, Robert C. Robinson2-4, Daiwen Yang*,1
1 Department of Biological Sciences, National University of Singapore, 14 Science Drive 4, Singapore 117543
2 Institute of Molecular and Cell Biology, A*STAR (Agency for Science, Technology and Research), Singapore
3 Department of Biochemistry, National University of Singapore, Singapore.
4 NTU Institute of Structural Biology, Nanyang Technological University, 59 Nanyang Drive, Singapore 636921.
Published online in Scientific Reports on 28 March 2017.
Please see http://www.nature.com/articles/srep45230
Six-domain gelsolin regulates actin structural dynamics through its abilities to sever, cap and uncap F-actin. These activities are modulated by various cellular parameters like Ca2+ and pH. Until now, only the molecular activation mechanism of gelsolin by Ca2+ has been understood relatively well. The fragment comprising the first domain and six residues from the linker region into the second domain has been shown to be similar to the full-length protein in F-actin severing activity in the absence of Ca2+ at pH 5. To understand how this gelsolin fragment is activated for F-actin severing by lowering pH, we solved its NMR structures at both pH 7.3 and 5 in the absence of Ca2+ and measured the pKa values of acidic amino acid residues and histidine residues. The overall structure and dynamics of the fragment are not affected significantly by pH. Nevertheless, local structural changes caused by protonation of His29 and Asp109 results in the activation on lowering the pH, and protonation of His151 directly effects filament binding since it resides in the gelsolin/actin interface. Mutagenesis studies support that His29, Asp109 and His151 play important roles in the pH-dependent severing activity of the gelsolin fragment.
Location of critical gelsolin residues in relation to F-actin binding. A) The structure of gelsolin domain 1 (G1+) bound to G-actin was superimposed onto a protomer in the structure of F-actin. B) A close up of the gelsolin/actin interface. Only one F-actin protomer (green) is shown together with the G-actin (grey). G1+ is painted yellow with regions colored in cyan showing G1+ interactions within the model which have no steric clash with F-actin, whereas regions highlighted in pink have steric clashes highlighted by the red ellipse. Ca2+ ions are indicated by blue balls. Residues that contribute to pH-dependent actin severing are labelled.
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