Article
Received: 14 November 2008; Revised: 12 December 2008; Accepted: 17 December 2008
10.1002/pro.59 About DOI
Dematin exhibits a natively unfolded core domain and an independently folded headpiece domain |
| Lin Chen 1, Zhenghui G. Jiang 1, Anwar A. Khan 2, Athar H. Chishti 2, C. James McKnight 1 * |
| 1Department of Physiology and Biophysics, Boston University School of Medicine, Boston, Massachusetts 02118 2Department of Pharmacology and Cancer Center, University of Illinois College of Medicine, Chicago, Illinois 60612 |
| email: C. James McKnight (cjmck@bu.edu) |
*Correspondence to C. James McKnight, 700 Albany Street, W407, Boston, MA 02118
Funded by:
National Institutes of Health; Grant Number: GM62886, HL051445
| Keywords |
| erythrocyte dematin band 4.9 PEST sequence NMR headpiece domain natively unfolded protein actin binding protein |
| Abstract |
Dematin is an actin-binding protein originally identified in the junctional complex of the erythrocyte plasma membrane, and is present in many nonerythroid cells. Dematin headpiece knockout mice display a spherical red cell phenotype and develop a compensated anemia. Dematin has two domains: a 315-residue, proline-rich  core domain and a 68-residue carboxyl-terminal villin-type headpiece domain. Expression of full-length dematin in E. coli as a GST recombinant protein results in truncation within a proline, glutamic acid, serine, threonine rich region (PEST). Therefore, we designed a mutant construct that replaces the PEST sequence. The modified dematin has high actin binding activity as determined by actin sedimentation assays. Negative stain electron microscopy demonstrates that the modified dematin also exhibits actin bundling activity like that of native dematin. Circular dichroism (CD) and NMR spectral analysis, however, show little secondary structure in the modified dematin. The lack of secondary structure is also observed in native dematin purified from human red blood cells. 15N-HSQC NMR spectra of modified dematin indicate that the headpiece domain is fully folded whereas the core region is primarily unfolded. Our finding suggests that the core is natively unfolded and may serve as a scaffold to organize the components of the junctional complex. |
Received: 14 November 2008; Revised: 12 December 2008; Accepted: 17 December 2008
| Digital Object Identifier (DOI) |
10.1002/pro.59 About DOI
